Formulation of dual cycloxygenase (COX) and lipoxygenase (LOX) inhibitors for mammal skin care

ABSTRACT

The present invention provides a novel composition of matter comprised of a mixture of two specific classes of compounds—Free-B-Ring flavonoids and flavans—for use in the prevention and treatment of diseases and conditions associated with the skin. This composition of matter simultaneously inhibits cyclooxygenase (COX) and lipoxygenase (LOX) enzymatic activity in normal, aged and damaged dermal cells and tissues. This invention further provides a method for the prevention and treatment of diseases and conditions of the skin mediated by cyclooxygenase (COX) and lipoxygenase (LOX). The method for preventing and treating COX and LOX mediated diseases and conditions of the skin is comprised of topically administering to a host in need thereof a therapeutically effective amount of a composition comprising a mixture of Free-B-Ring flavonoids and flavans synthesized and/or isolated from a single plant or multiple plants, preferably in the  Scutellaria  and  Acacia  genus of plants and pharmaceutically and/or cosmetically acceptable carriers. Finally the present invention provides a method for the prevention and treatment of COX and LOX mediated diseases and conditions, including but not limited to sun burns, thermal burns, acne, topical wounds, minor inflammatory conditions caused by fungal, microbial and viral infections, vitilago, systemic lupus erythromatosus, psoriasis, carcinoma, melanoma, as well as other mammal skin cancers, skin damage resulting from exposure to ultraviolet (UV) radiation, chemicals, heat, wind and dry environments, wrinkles, saggy skin, lines and dark circles around the eyes, dermatitis and other allergy related conditions of the skin. Use of the composition described herein also affords the benefit of smooth and youthful skin with improved elasticity, reduced and delayed aging, enhanced youthful appearance and texture, and increased flexibility, firmness, smoothness and suppleness.

FIELD OF THE INVENTION

[0001] This invention relates generally to a method for the preventionand treatment of diseases and conditions mediated by cyclooxygenase(COX) and lipoxygenase (LOX). Specifically, the present inventionrelates to a novel composition of matter comprised of a mixture of ablend of two specific classes of compounds—Free-B-Ring flavonoids andflavans—for use in the prevention and treatment of diseases andconditions of the skin mediated by the COX and LOX pathways. Included inthe present invention is a method for the prevention and treatment ofCOX and LOX mediated diseases and conditions, including but not limitedto sun burns, thermal burns, acne, topical wounds, minor inflammatoryconditions caused by fungal, microbial and viral infections, vitilago,systemic lupus erythromatosus, psoriasis, carcinoma, melanoma, as wellas other mammal skin cancers, skin damage resulting from exposure toultraviolet (UV) radiation, chemicals, heat, wind and dry environments,wrinkles, saggy skin, lines and dark circles around the eyes, dermatitisand other allergy related conditions of the skin. Use of the compositiondescribed herein also affords the benefit of smooth and youthful skinwith improved elasticity, reduced and delayed aging, enhanced youthfulappearance and texture, and increased flexibility, firmness, smoothnessand suppleness.

BACKGROUND OF THE INVENTION

[0002] Sunlight has a significant effect on the skin causing prematureaging, skin cancer and a host of other skin changes such as erythema andtanning. The majority of the damage caused by sunlight is attributed toultraviolet (UV) radiation, which has a wavelength from 200 nm to 400nm. Ultraviolet radiation is divided into three categories, UVA, UVB orUVC, depending on wavelength. UVA, which has a wavelength range from320-400 nm, can cause tanning and mild sunburn. UVB, which has awavelength range from 290-320 nm, can cause sunburn and stimulatepigmentation. UVC, which has a wavelength range from 100-290 nm, cancause damage but not tanning. Exposure of the skin to UV radiationinduces biphasic reactions. Thus, upon initial exposure an immediateerythema reaction occurs, which is a weak reaction that fades within 30minutes. A delayed erythema reaction occurs after 2-5 hours of exposureand peaks around 10-24 hours. Enhanced prostaglandin and leukotrieneproduction are the major mechanisms of action for UV, sun andchemical/thermal caused erythema. (Wang (2002) Adv. Dermatol. 18:247).

[0003] The liberation and metabolism of arachidonic acid (AA) from thecell membrane results in the generation of pro-inflammatory metabolitesby several different pathways. Arguably, two of the most importantpathways to inflammation are mediated by the enzymes lipoxygenase (LOX)and cyclooxygenase (COX). These are parallel pathways that result in thegeneration of leukotrienes and prostaglandins, respectively, which playimportant roles in the initiation and progression of the inflammatoryresponse. These vasoactive compounds are chemotaxins, which both promoteinfiltration of inflammatory cells into tissues and serve to prolong theinflammatory response. Consequently, the enzymes responsible forgenerating these mediators of inflammation have become the targets inthe current invention to develop topically administered therapeuticagents aimed at the dual inhibition of inflammation resulting from bothpathways which contribute to the physiological and pathologicalprocesses of diseases and conditions such as sun burn, thermal burns,scald, acne, topical wounds, lupus erythromatosus, psoriasis, carcinoma,melanoma, and other mammalian skin cancers.

[0004] Inhibition of the COX enzyme is the mechanism of actionattributed to most non-steroidal anti-inflammatory drugs (NSAIDS). Thereare two distinct isoforms of the COX enzyme (COX-1 and COX-2), whichshare approximately 60% sequence homology, but differ in expressionprofiles and function. COX-1 is a constitutive form of the enzyme thathas been linked to the production of physiologically importantprostaglandins, which help regulate normal physiological functions, suchas platelet aggregation, protection of cell function in the stomach andmaintenance of normal kidney function. (Dannhardt and Kiefer (2001) Eur.J. Med. Chem. 36:109-26). The second isoform, COX-2, is a form of theenzyme that is inducible by pro-inflammatory cytokines, such asinterleukin-1β (IL-1β) and other growth factors. (Herschmann (1994)Cancer Metastasis Rev. 134:241-56; Xie et al. (1992) Drugs Dev. Res.25:249-65). This isoform catalyzes the production of prostaglandin E₂(PGE2) from arachidonic acid (AA). Inhibition of COX is responsible forthe anti-inflammatory activity of conventional NSAIDs.

[0005] Inhibitors that demonstrate dual specificity for COX and LOXwould have the obvious benefit of inhibiting multiple pathways ofarachidonic acid metabolism. Such inhibitors would block theinflammatory effects of prostaglandins (PG), as well as, those ofmultiple leukotrienes (LT) by limiting their production. This includesthe vasodilation, vasopermeability and chemotactic effects of PGE2,LTB4, LTD4 and LTE4, also known as the slow reacting substance ofanaphalaxis. Of these, LTB4 has the most potent chemotactic andchemokinetic effects. (Moore (1985) in Prostanoids: pharmacological,physiological and clinical relevance, Cambridge University Press, N.Y.,pp. 229-230).

[0006] Because the mechanism of action of COX inhibitors overlaps thatof most conventional NSAID's, COX inhibitors are used to treat many ofthe same symptoms, including pain and swelling associated withinflammation in transient conditions and chronic diseases in whichinflammation plays a critical role. Transient conditions includetreatment of inflammation associated with minor abrasions or contactdermatitis, as well as, skin conditions that are directly associatedwith the prostaglandin and leukotriene pathways, such as skinhyperpigmentation, age spots, vitilago, systemic lupus erythromatosus,psoriasis, carcinoma, melanoma, and other mammalian skin cancers. Theuse of COX inhibitors has been expanded to include diseases, such assystemic lupus erythromatosus (SLE) (Goebel et al. (1999) Chem. Res.Toxicol. 12:488-500; Patrono et al. (1985) J. Clin. Invest.76:1011-1018), as well as, rheumatic skin conditions, such asscleroderma. COX inhibitors are also used for the relief of inflammatoryskin conditions that are not of rheumatic origin, such as psoriasis, inwhich reducing the inflammation resulting from the overproduction ofprostaglandins could provide a direct benefit. (Fogh et al. (1993) ActaDerm Venerologica 73:191-193). Recently over expression of5-lipoxygenase in the skin of patients with system sclerosis has beenreported. This has led to the suggestion that the LOX pathway may be ofsignificance in the pathogenesis of system sclerosis and may represent avalid therapeutic target. (Kowal-Bielecka (2001) Arthritis Rheum.44(8):1865). Finally, the increased enzymatic activity of both the COX-2and 5-LOX at the site of allergen injections suggests the potential forusing dual COX/LOX inhibitors to treat the symptoms of both the earlyand late phases of the skin allergic response. (Church (2002) Clin. Exp.Allergy. 32(7):1013).

[0007] Topical application of a selective cyclooxygenase inhibitor hasbeen shown to suppress UVB mediated cutaneous inflammation followingboth acute and long-term exposure. Additionally, edema, dermalneutrophil infiltration and activation, PGE2 levels and the formation ofsunburn cells were reduced by the topical application of a COXinhibitor. (Wilgus (2000) Prostaglandins Other Lipid Mediat. 62(4):367).The COX inhibitor Celebrex™ has been shown to reduce the effects of UVinduced inflammation when administered systematically (Wilgus et al.(2002) Adv. Exp. Med. Biol. 507:85), and topically (Wilgus et al. (2000)Protaglandins Other Lipid Mediat. 62:367). In animal models, the knownCOX inhibitor aspirin and various lipoxygenase inhibitors exhibitedvasoprotective activity against inflammation and vasodepressionresulting from UV irradiation. (Kuhn (1988) Biomed. Biochim. Acta.47:S320). Acute or long-term chronic UV exposure causes skin damage andphotoageing that are characterized by degradation of collagen andaccumulation of abnormal elastin in the superficial dermis. A dualCOX/LOX inhibitor can be utilized to prevent and treat collagendegradation caused by inflammatory infiltration by significantlyreducing the vasodilating, vasopermeability, chemotactic andchemotaxins-prostaglandins (PG), as well as, those of multipleleukotrienes (LT). (Bosset (2003) Br. J. Dermatol. 149(4):826; Hase(2000) Br. J. Dermatol. 142(2):267). Additionally, chemically inducedoxidative stress in mouth skin can be inhibited by separatelyadministrating COX and LOX inhibitors to reduce leukocyte adhesion,infiltration and H₂O₂ generation. (Nakamura (2003) Free Radical Biol.Med. 35(9):997).

[0008] In addition to their use as anti-inflammatory agents, anotherpotential role for COX inhibitors is in the treatment of cancer. Overexpression of COX has been demonstrated in various human malignanciesand inhibitors of COX have been shown to be efficacious in the treatmentof animals with skin tumors. While the mechanism of action is notcompletely understood, the over expression of COX has been shown toinhibit apoptosis and increase the invasiveness of tumorgenic celltypes. (Dempke et al. (2001) J. Can. Res. Clin. Oncol. 127:411-17; Mooreand Simmons (2000) Current Med. Chem. 7:1131-1144). Up regulated COXproduction has been implicated in the generation of actinic keratosisand squamous cell carcinoma in skin. Increased amounts of COX were alsofound in lesions produced by DNA damage. (Buckman et al. (1998)Carcinogenesis 19:723). Therefore, control of expression or proteinfunction of COX would seem to lead to a decrease in the inflammatoryresponse and the eventual progression to cancer. In fact, COX inhibitorssuch as indomethacin and Celebrex™ have been found to be effective intreating UV induced erythema and tumor formation. (Fischer (1999) Mol.Carcinog. 25:231; Pentland (1999) Carcinogenesis 20:1939). Recently, theover expression of lipoxygenase has also been shown to be related toepidermal tumor development (Muller (2002) Cancer Res. 62(16):4610) andmelanoma carcinogenesis (Winer (2002) Melanoma Res. 12(5):429). Thearachidonic acid (AA) metabolites generated from lipoxygenase pathwaysplay important roles in tumor growth related signal transductionsuggesting that that the inhibition of lipoxygenase pathways should be avalid target to prevent cancer progression. (Cuendet (2000) Drug MetabolDrug Interact 17(4): 109; Steele (2003) Mutat Res. 523-524:137). Thus,the use of therapeutic agents having dual COX/LOX inhibitory activityoffers significant advantages in the chemoprevention of cancer.

[0009] Prostaglandins and leukotrienes also play important roles in thephysiological and pathological processes of wounds, burns, scald, acne,microbial infections, dermatitis, and many other diseases and conditionsof the skin. The activation of a pro-inflammatory cascade after thermalor chemical burns with significantly elevated cyclooxygenase andlipoxygenase activities are well documented and play an important rolein the development of subsequent severe symptoms and immune dysfunctionthat may lead to multiple organ failure. (Schwacha (2003) Burns 29(1):1;He (2001) J. Bum Care Rehabil. 22(1):58).

[0010] Acne is a disease of the pilosebaceous unit with abnormalities insebum production, follicular epithelial desquamation, bacterialproliferation and inflammation. The inflammatory properties of acne canbe detected by polarized light photography and utilized for clinicaldiagnosis, including an evaluation of the extent of the acne and also todetermine the effectiveness of therapy. (Phillips (1997) J. Am. Acad.Dermatol. 37(6):948). Current therapeutic agents for the prevention andtreatment of acne include anti-inflammatory agents, like retinoids,antimicrobial agents and hormonal drugs. (Leyden (2003) J. Am. Acad.Dermatol. 49(3 Suppl):S200). Topical application of anti-inflammatorydrugs, such as retinoids (Millikan (2003) J. Am. Acad. Dermatol.4(2):75) and the COX inhibitor salicylic acid (Lee (2003) Dermnatol Surg29(12):1196) have been clinically demonstrated as an effective and safetherapy for the treatment of acne. Additionally, the use of nonsteroidalanti-inflammatory drugs (NSAIDs) are well documented as therapeuticagents for common and uncommon dermatoses, including acne, psoriasis,sun burn, erythema nodosum, cryoglobulinemia, Sweet's syndrome, systemicmastocytosis, urticarial, liverdoid and nodular vasculitis. (Friedman(2002) J. Cutan Med. Surg. 6(5):449).

[0011] Flavonoids or bioflavonoids are a widely distributed group ofnatural products, which have been reported to have antibacterial,anti-inflammatory, antiallergic, antimutagenic, antiviral,antineoplastic, anti-thrombic and vasodilatory activity. The structuralunit common to this group of compounds includes two benzene rings oneither side of a 3-carbon ring as illustrated by the following generalstructural formula:

[0012] Various combinations of hydroxyl groups, sugars, oxygen andmethyl groups attached to this general three ring structure create thevarious classes of flavonoids, which include flavanols, flavones,flavan-3-ols (catechins), anthocyanins and isoflavones.

[0013] Free-B-Ring flavones and flavonols are a specific class offlavonoids, which have no substituent groups on the aromatic B ring(referred to herein as Free-B-Ring flavonoids), as illustrated by thefollowing general structure:

[0014] wherein

[0015] R₁, R₂, R₃, R_(4,) and R₅ are independently selected from thegroup consisting of —H, —OH, —SH, OR, —SR, —NH₂, —NHR, —NR₂, —NR₃ ⁺X⁻, acarbon, oxygen, nitrogen or sulfur, glycoside of a single or acombination of multiple sugars including, but not limited toaldopentoses, methyl-aldopentose, aldohexoses, ketohexose and theirchemical derivatives thereof;

[0016] wherein

[0017] R is an alkyl group having between 1-10 carbon atoms; and

[0018] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,fluoride, sulfate, phosphate, acetate, carbonate, etc.

[0019] Free-B-Ring flavonoids are relatively rare. Out of 9,396flavonoids synthesized or isolated from natural sources, only 231Free-B-Ring flavonoids are known (The Combined Chemical Dictionary,Chapman & Hall/CRC, Version 5:1 June 2001). Free-B-Ring flavonoids havebeen reported to have diverse biological activity. For example, galangin(3,5,7-trihydroxyflavone) acts as antioxidant and free radical scavengerand is believed to be a promising candidate for anti-genotoxicity andcancer chemoprevention. (Heo et al. (2001) Mutat. Res. 488(2):135-150).It is an inhibitor of tyrosinase monophenolase (Kubo et al. (2000)Bioorg. Med. Chem. 8(7):1749-1755), an inhibitor of rabbit heartcarbonyl reductase (Imamura et al. (2000) J. Biochem. 127(4):653-658),has antimicrobial activity (Afolayan and Meyer (1997) Ethnopharmacol.57(3):177-181) and antiviral activity (Meyer et al. (1997) J.Ethnopharmacol. 56(2):165-169). Baicalein and two other Free-B-Ringflavonoids, have antiproliferative activity against human breast cancercells. (So et al. (1997) Cancer Lett. 112(2):127-133).

[0020] Typically, flavonoids have been tested for biological activityrandomly based upon their availability. Occasionally, the requirement ofsubstitution on the B-ring has been emphasized for specific biologicalactivity, such as the B-ring substitution required for high affinitybinding to p-glycoprotein (Boumendjel et al. (2001) Bioorg. Med. Chem.Lett. 11(1):75-77); cardiotonic effect (Itoigawa et al. (1999) J.Ethnopharmacol. 65(3): 267-272), protective effect on endothelial cellsagainst linoleic acid hydroperoxide-induced toxicity (Kaneko and Baba(1999) Biosci Biotechnol. Biochem 63(2):323-328), COX-1 inhibitoryactivity (Wang (2000) Phytomedicine 7:15-19) and prostaglandinendoperoxide synthase (Kalkbrenner et al. (1992) Pharmacology44(1):1-12). Only a few publications have mentioned the significance ofthe unsubstituted B ring of the Free-B-Ring flavonoids. One example, isthe use of 2-phenyl flavones, which inhibit NADPH quinone acceptoroxidoreductase, as potential anticoagulants. (Chen et al. (2001)Biochem. Pharmacol. 61(11):1417-1427).

[0021] The mechanism of action with respect to the anti-inflammatoryactivity of various Free-B-Ring flavonoids has been controversial. Theanti-inflammatory activity of the Free-B-Ring flavonoids, chrysin (Lianget al. (2001) FEBS Lett. 496(1):12-18), wogonin (Chi et al. (2001)Biochem. Pharmacol. 61:1195-1203) and halangin (Raso et al. (2001) LifeSci. 68 :921-93 1), has been associated with the suppression ofinducible cyclooxygenase and nitric oxide synthase via activation ofperoxisome proliferator activated receptor gamma (PPARγ) and influenceon degranulation and AA release. (Tordera et al. (1994) Z. Naturforsch[C] 49:235-240). It has been reported that oroxylin, baicalein andwogonin inhibit 12-lipoxygenase activity without affectingcyclooxygenase. (You et al. (1999) Arch. Pharm. Res. 22(1):18-24). Morerecently, the anti-inflammatory activity of wogonin, baicalin andbaicalein has been reported as occurring through inhibition of induciblenitric oxide synthase and cox-2 gene expression induced by nitric oxideinhibitors and lipopolysaccharide. (Chen et al. (2001) Biochem.Pharmacol. 61(11):1417-1427). It has also been reported that oroxylinacts via suppression of NFκB activation. (Chen et al. (2001) Biochem.Pharmacol. 61(11):1417-1427). Finally, wogonin reportedly inhibitsinducible PGE2 production in macrophages. (Wakabayashi and Yasui (2000)Eur. J. Pharmacol. 406(3):477-481).

[0022] Inhibition of the phosphorylation of mitrogen-activated proteinkinase and inhibition of Ca²⁺ ionophore A23187 induced PGE₂ release bybaicalein has been reported as the mechanism of anti-inflammatoryactivity of Scutellariae radix. (Nakahata et al. (1999) NipponYakurigaku Zasshi, 114, Supp. 11:215P-219P; Nakahata et al. (1998) Am.J. Chin Med. 26:311-323). Baicalin from Scutellaria baicalensis,reportedly inhibits superantigenic staphylococcal exotoxins stimulatedT-cell proliferation and production of IL-1β, IL-6, tumor necrosisfactor-α (TNF-α), and interferon-γ (IFN-γ). (Krakauer et al. (2001) FEBSLett. 500:52-55). Thus, the anti-inflammatory activity of baicalin hasbeen associated with inhibiting the pro-inflammatory cytokines mediatedsignaling pathways activated by superantigens. However, it has also beenproposed that the anti-inflammatory activity of baicalin is due to thebinding of a variety of chemokines, which limits their biologicalactivity. (Li et al. (2000) Immunopharmacology 49:295-306). Recently,the effects of baicalin on adhesion molecule expression induced bythrombin and thrombin receptor agonist peptide (Kimura et al. (2001)Planta Med. 67:331-334), as well as, the inhibition of mitogen-activatedprotein kinase cascade (MAPK) (Nakahata et al. (1999) Nippon YakurigakuZasshi, 114, Supp 11:215P-219P; Nakahata et al. (1998) Am. J. Chin Med.26:311-323) have been reported.

[0023] The Chinese medicinal plant, Scutellaria baicalensis containssignificant amounts of Free-B-Ring flavonoids, including baicalein,baicalin, wogonin and baicalenoside. Traditionally, this plant has beenused to treat a number of conditions including clearing away heat,purging fire, dampness-warm and summer fever syndromes; polydipsiaresulting from high fever; carbuncle, sores and other pyogenic skininfections; upper respiratory infections, such as acute tonsillitis,laryngopharyngitis and scarlet fever; viral hepatitis; nephritis;pelvitis; dysentery; hematemesis and epistaxis. This plant has alsotraditionally been used to prevent miscarriage. (Encyclopedia of ChineseTraditional Medicine, ShangHai Science and Technology Press, ShangHai,China, 1998). Clinically Scutellaria is now used to treat conditionssuch as pediatric pneumonia, pediatric bacterial diarrhea, viralhepatitis, acute gallbladder inflammation, hypertension, topical acuteinflammation, resulting from cuts and surgery, bronchial asthma andupper respiratory infections. (Encyclopedia of Chinese TraditionalMedicine, ShangHai Science and Technology Press, ShangHai, China, 1998).The pharmacological efficacy of Scutellaria roots for treating bronchialasthma is reportedly related to the presence of Free-B-Ring flavonoidsand their suppression of eotaxin associated recruitment of eosinophils.(Nakajima et al. (2001) Planta Med. 67(2):132-135).

[0024] To date, a number of naturally occurring Free-B-Ring flavonoidshave been commercialized for varying uses. For example, liposomeformulations of Scutellaria extracts have been utilized for skin care(U.S. Pat. Nos. 5,643,598; 5,443,983). Baicalin has been used forpreventing cancer, due to its inhibitory effects on oncogenes (U.S. Pat.No. 6,290,995). Baicalin and other compounds have been used asantiviral, antibacterial and immunomodulating agents (U.S. Pat. No.6,083,921 and WO98/42363) and as natural anti-oxidants (WO98/49256 andPoland Pub. No. 9,849,256). Scutellaria baicalensis root extract hasbeen formulated as a supplemental sun screen agent with additive effectsof the cumulative SPFs of each individual component in a topicalformulation (WO98/19651). Chrysin has been used for its anxiety reducingproperties (U.S. Pat. No. 5,756,538). Anti-inflammatory flavonoids areused for the control and treatment of anorectal and colonic diseases(U.S. Pat. No. 5,858,371), and inhibition of lipoxygenase (U.S. Pat. No.6,217,875). These compounds are also formulated with glucosaminecollagen and other ingredients for repair and maintenance of connectivetissue (U.S. Pat. No. 6,333,304). Flavonoid esters constitute activeingredients for cosmetic compositions (U.S. Pat. No. 6,235,294). U.S.application Ser. No. 10/091,362, filed Mar. 1, 2002, entitled“Identification of Free-B-Ring Flavonoids as Potent COX-2 Inhibitors,”and U.S. application Ser. No. 10/427,746, filed Jul. 22, 2003, entitled“Formulation of a Mixture of Free-B-Ring Flavonoids and Flavans as aTherapeutic Agent” both disclose a method for inhibiting thecyclooxygenase enzyme COX-2 by administering a composition comprising aFree-B-Ring flavonoid or a composition containing a mixture ofFree-B-Ring flavonoids to a host in need thereof. This is the firstreport of a link between Free-B-Ring flavonoids and COX-2 inhibitoryactivity. These applications are specifically incorporated herein byreference in their entirety.

[0025] Japanese Pat. No. 63027435, describes the extraction, andenrichment of baicalein and Japanese Pat. No. 61050921 describes thepurification of baicalin.

[0026] Flavans include compounds illustrated by the following generalstructure:

[0027] wherein

[0028] R₁, R₂, R₃, R₄ and R₅ are independently selected from the groupconsisting of —H, —OH, —SH, —OCH₃, —SCH₃, —OR, —SR, —NH₂, —NRH, —NR₂,—NR₃ ⁺X⁻, esters of the mentioned substitution groups, including, butnot limited to, gallate, acetate, cinnamoyl and hydroxyl-cinnamoylesters, trihydroxybenzoyl esters and caffeoyl esters, and their chemicalderivatives thereof; a carbon, oxygen, nitrogen or sulfur glycoside of asingle or a combination of multiple sugars including, but not limitedto, aldopentoses, methyl aldopentose, aldohexoses, ketohexose and theirchemical derivatives thereof; dimer, trimer and other polymerizedflavans;

[0029] wherein

[0030] R is an alkyl group having between 1-10 carbon atoms; and

[0031] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride, and carbonate, etc.

[0032] Catechin is a flavan, found primarily in green tea, having thefollowing structure:

[0033] Catechin works both alone and in conjunction with otherflavonoids found in tea, and has both antiviral and antioxidantactivity. Catechin has been shown to be effective in the treatment ofviral hepatitis. It also appears to prevent oxidative damage to theheart, kidney, lungs and spleen and has been shown to inhibit the growthof stomach cancer cells.

[0034] Catechin and its isomer epicatechin inhibit prostaglandinendoperoxide synthase with an IC₅₀ value of 40 μM. (Kalkbrenner et al.(1992) Pharmacol. 44:1-12). Five flavan-3-ol derivatives, including(+)-catechin and gallocatechin, isolated from four plant species: Atunaracemosa, Syzygium carynocarpum, Syzygium malaccense and Vantaneaperuviana, exhibit equal to weaker inhibitory activity against COX-2,relative to COX-1, with IC₅₀ values ranging from 3.3 μM to 138 μM(Noreen et al. (1998) Planta Med. 64:520-524). (+)-Catechin, isolatedfrom the bark of Ceiba pentandra, inhibits COX-1 with an IC₅₀ value of80 μM (Noreen et al. (1998) J. Nat. Prod. 61:8-12). Commerciallyavailable pure (+)-catechin inhibits COX-1 with an IC₅₀ value of around183 to 279 μM depending upon the experimental conditions, with noselectivity for COX-2. (Noreen et al. (1998) J. Nat. Prod. 61:1-7).

[0035] Green tea catechin, when supplemented into the diets of Spraguedawley male rats, lowered the activity level of platelet PLA₂ andsignificantly reduced platelet cyclooxygenase levels. (Yang et al.(1999) J. Nutr. Sci. Vitaminol. 45:337-346). Catechin and epicatechinreportedly weakly suppress cox-2 gene transcription in human coloncancer DLD-1 cells (IC₅₀=415.3 μM). (Mutoh et al. (2000) Jpn. J. CancerRes. 91:686-691). The neuroprotective ability of (+)-catechin from redwine results from the antioxidant properties of catechin, rather thaninhibitory effects on intracellular enzymes, such as cyclooxygenase,lipoxygenase, or nitric oxide synthase (Bastianetto et al. (2000) Br. J.Pharmacol. 131:711-720). Catechin derivatives purified from green andblack tea, such as epigallocatechin-3-gallate (EGCG), epigallocatechin(EGC), epicatechin-3-gallate (ECG), and theaflavins showed inhibition ofcyclooxygenase and lipoxygenase dependent metabolism of AA in humancolon mucosa and colon tumor tissues (Hong et al. (2001) Biochem.Pharmacol. 62:1175-1183) and induce cox-2 expression and PGE₂ production(Park et al. (2001) Biochem. Biophys. Res. Commun. 286:721-725).Epiafzelechin isolated from the aerial parts of Celastrus orbiculatusexhibited dose-dependent inhibition of COX-1 activity with an IC₅₀ valueof 15 μM and also demonstrated anti-inflammatory activity againstcarrageenin-induced mouse paw edema following oral administration at adosage of 100 mg/kg. (Min et al. (1999) Planta Med. 65:460-462).

[0036]Acacia is a genus of leguminous trees and shrubs. The genus Acaciaincludes more than 1000 species belonging to the family of Leguminosaeand the subfamily of Mimosoideae. Acacias are distributed worldwide intropical and subtropical areas of Central and South America, Africa,parts of Asia, as well as, Australia, which has the largest number ofendemic species. Acacias are very important economically, providing asource of tannins, gums, timber, fuel and fodder. Tannins, which areisolated primarily from bark, are used extensively for tanning hides andskins. Some Acacia barks are also used for flavoring local spirits. Someindigenous species like A. sinuata also yield saponins, which are any ofvarious plant glucosides that form soapy lathers when mixed and agitatedwith water. Saponins are used in detergents, foaming agents andemulsifiers. The flowers of some Acacia species are fragrant and used tomake perfume. The heartwood of many Acacias is used for makingagricultural implements and also provides a source of firewood. Acaciagums find extensive use in medicine and confectionary and as sizing andfinishing materials in the textile industry.

[0037] To date, approximately 330 compounds have been isolated fromvarious Acacia species. Flavonoids are the major class of compoundsisolated from Acacias. Approximately 180 different flavonoids have beenidentified, 111 of which are flavans. Terpenoids are second largestclass of compounds isolated from species of the Acacia genus, with 48compounds having been identified. Other classes of compounds isolatedfrom Acacia include, alkaloids (28), amino acids/peptides (20), tannins(16), carbohydrates (15), oxygen heterocycles (15) and aliphaticcompounds (10). (Buckingham, The Combined Chemical Dictionary, Chapman &Hall CRC, version 5:2, December 2001).

[0038] Phenolic compounds, particularly flavans are found in moderate tohigh concentrations in all Acacia species. (Abdulrazak et al. (2000)Journal of Animal Sciences. 13:935-940). Historically, most of theplants and extracts of the Acacia genus have been utilized asastringents to treat gastrointestinal disorders, diarrhea, indigestionand to stop bleeding. (Vautrin (1996) Universite Bourgogne (France)European abstract 58-01C:177; Saleem et al. (1998) Hamdard Midicus.41:63-67). The bark and pods of Acacia arabica Willd. contain largequantities of tannins and have been utilized as astringents andexpectorants. (Nadkarni (1996) India Materia Medica, Bombay PopularPrakashan, pp. 9-17). Diarylpropanol derivatives, isolated from stembark of Acacia tortilis from Somalia, have been reported to have smoothmuscle relaxing effects. (Hagos et al. (1987) Planta Medica. 53:27-31,1987). It has also been reported that terpenoid saponins isolated fromAcacia victoriae have an inhibitory effect ondimethylbenz(a)anthracene-induced murine skin carcinogenesis (Hanauseket al. (2000) Proceedings American Association for Cancer ResearchAnnual Meeting 41:663) and induce apotosis (Haridas et al. (2000)Proceedings American Association for Cancer Research Annual Meeting.41:600). Plant extracts from Acacia nilotica have been reported to havespasmogenic, vasoconstrictor and anti-hypertensive activity (Amos et al.(1999) Phytotherapy Research 13:683-685; Gilani et al. (1999)Phytotherapy Research. 13:665-669), and antiplatelet aggregatoryactivity (Shah et al. (1997) General Pharmacology. 29:251-255).Anti-inflammatory activity has been reported for A. nilotica. It wasspeculated that flavonoids, polysaccharides and organic acids werepotential active components. (Dafallah and Al-Mustafa (1996) AmericanJournal of Chinese Medicine. 24:263-269). To date, the only reported5-lipoxygenase inhibitor isolated from Acacia is a monoterpenoidalcarboxamide (Seikine et al. (1997) Chemical and Pharmaceutical Bulletin.45:148-11).

[0039] The extract from the bark of Acacia has been patented in Japanfor external use as a whitening agent (Abe, JP10025238), as a glucosyltransferase inhibitor for dental applications (Abe, JP07242555), as aprotein synthesis inhibitor (Fukai, JP 07165598), as an active oxygenscavenging agent for external skin preparations (Honda, JP 07017847,Bindra U.S. Pat. No. 6,1266,950), and as a hyaluronidase inhibitor fororal consumption to prevent inflammation, pollinosis and cough (Ogura,JP 07010768).

[0040] To date, Applicant is unaware of any reports of a formulationcombining only Free-B-Ring-Flavonoids and flavans as the primarybiologically active components for the dual inhibition of the COX/LOXenzymes that yield significant benefit to mammal skin conditions.

SUMMARY OF THE INVENTION

[0041] The present invention includes methods that are effective insimultaneously inhibiting both the cyclooxygenase (COX) and lipoxygenase(LOX) enzymes, for use in the prevention and treatment of diseases andconditions related to the skin. The method for the simultaneous dualinhibition of the COX and LOX enzymes is comprised administering,preferably topically, a composition comprised of a mixture ofFree-B-Ring flavonoids and flavans synthesized and/or isolated from asingle plant or multiple plants to a host in need thereof. Thiscomposition of matter is referred to herein as Soliprin™. The efficacyof this method was demonstrated with purified enzymes, in different celllines, in multiple animal models and eventually in a human clinicalstudy. The ratio of the Free-B-Ring flavonoids to flavans in thecomposition can be in the range of 99.9:0.1 of Free-B-Ringflavonoids:flavans to 0.1:99.9 Free-B-Ring flavonoids:flavans. Inspecific embodiments of the present invention, the ratio of Free-B-Ringflavonoids to flavans is selected from the group consisting ofapproximately 90:10, 80:20, 70:30, 60:40, 50:50, 40:60, 30:70, 20:80 and10:90. In a preferred embodiment of this invention, the ratio ofFree-B-Ring flavonoids:flavans in the composition of matter is 80:20. Ina preferred embodiment, the Free-B-Ring flavonoids are isolated from aplant or plants in the Scutellaria genus of plants and the flavans areisolated from a plant or plants in the Acacia genus of plants.

[0042] The present invention also includes methods for the preventionand treatment of COX and LOX mediated diseases and conditions of theskin. The method for preventing and treating COX and LOX mediateddiseases and conditions of the skin is comprised of administering,preferably topically, to a host in need thereof an effective amount of acomposition comprising a mixture of Free-B-Ring flavonoids and flavanssynthesized and/or isolated from a single plant or multiple plants and apharmaceutically acceptable carrier. The ratio of the Free-B-Ringflavonoids to flavans in the composition can be in the range of 99.9:0.1of Free-B-Ring flavonoids:flavans to 0.1:99.9 Free-B-Ringflavonoids:flavans. In specific embodiments of the present invention,the ratio of Free-B-Ring flavonoids to flavans is selected from thegroup consisting of approximately 90: 10, 80:20, 70:30, 60:40, 50:50,40:60, 30:70, 20:80 and 10:90. In a preferred embodiment of thisinvention, the ratio of Free-B-Ring flavonoids:flavans in thecomposition of matter is 80:20. In a preferred embodiment, theFree-B-Ring flavonoids are isolated from a plant or plants in theScutellaria genus of plants and the flavans are isolated from a plant orplants in the Acacia genus of plants.

[0043] The Free-B-Ring flavonoids, also referred to herein asFree-B-Ring flavones and flavonols, that can be used in accordance withthe following invention include compounds illustrated by the followinggeneral structure:

[0044] wherein

[0045] R₁, R₂, R₃, R_(4,) and R₅ are independently selected from thegroup consisting of —H, —OH, —SH, OR, —SR, —NH₂, —NHR, —NR₂, —NR₃ ⁺X⁻, acarbon, oxygen, nitrogen or sulfur, glycoside of a single or acombination of multiple sugars including, but not limited toaldopentoses, methyl-aldopentose, aldohexoses, ketohexose and theirchemical derivatives thereof;

[0046] wherein

[0047] R is selected from an alkyl group having between 1-10 carbonatoms; and

[0048] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride, carbonate, etc.

[0049] The Free-B-Ring flavonoids of this invention may be obtained bysynthetic methods or extracted from the family of plants including, butnot limited to Annonaceae, Asteraceae, Bignoniaceae, Combretaceae,Compositae, Euphorbiaceae, Labiatae, Lauranceae, Leguminosae, Moraceae,Pinaceae, Pteridaceae, Sinopteridaceae, Ulmaceae and Zingiberacea. TheFree-B-Ring flavonoids can be extracted, concentrated, and purified fromthe following genus of high plants, including but not limited to Desmos,Achyrocline, Oroxylum, Buchenavia, Anaphalis, Cotula, Gnaphalium,Helichrysum, Centaurea, Eupatorium, Baccharis, Sapium, Scutellaria,Molsa, Colebrookea, Stachys, Origanum, Ziziphora, Lindera, Actinodaphne,Acacia, Derris, Glycyrrhiza, Millettia, Pongamia, Tephrosia, Artocarpus,Ficus, Pityrogramma, Notholaena, Pinus, Ulmus and Alpinia.

[0050] The flavans that can be used in accordance with the followinginvention include compounds illustrated by the following generalstructure:

[0051] wherein

[0052] R₁, R₂, R₃, R₄ and R₅ are independently selected from the groupconsisting of H, —OH, —SH, —OCH₃, —SCH₃, —OR, —SR, —NH₂, —NRH, —NR₂,—NR₃ ⁺X⁻, esters of the mentioned substitution groups, including, butnot limited to, gallate, acetate, cinnamoyl and hydroxyl-cinnamoylesters, trihydroxybenzoyl esters and caffeoyl esters; thereof carbon,oxygen, nitrogen or sulfur glycoside of a single or a combination ofmultiple sugars including, but not limited to, aldopentoses, methylaldopentose, aldohexoses, ketohexose and their chemical derivativesthereof; dimer, trimer and other polymerized flavans;

[0053] wherein

[0054] R is selected from an alkyl group having between 1-10 carbonatoms; and

[0055] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride, carbonate, etc.

[0056] The flavans of this invention may be obtained from a plant orplants selected from the genus of Acacia. In a preferred embodiment, theplant is selected from the group consisting of Acacia catechu, Acaciaconcinna, Acacia farnesiana, Acacia Senegal, Acacia speciosa, Acaciaarabica, A. caesia, A. pennata, A. sinuata. A. mearnsii, A. picnantha,A. dealbata, A. auriculiformis, A. holoserecia and A. mangium.

[0057] In one embodiment, the present invention includes a method forpreventing and treating a number of COX and LOX mediated diseases andconditions of the skin including, but not limited to sun burns, thermalburns, acne, topical wounds, minor inflammatory conditions caused byfungal, microbial and viral infections, vitilago, systemic lupuserythromatosus, psoriasis, carcinoma, melanoma, as well as other mammalskin cancers. In another embodiment the present invention includes amethod for preventing and treating skin damage resulting from exposureto ultraviolet (UV) radiation, chemicals, heat, wind and dryenvironments. In yet another embodiment the present invention includes amethod for preventing and treating wrinkles, saggy skin, lines and darkcircles around the eyes, dermatitis and other allergy related conditionsof the skin.

[0058] The present invention further includes therapeutic compositionscomprising the therapeutic agents of the present invention. In additionto their use for the prevention and treatment of the above describeddiseases and conditions of the skin, the therapeutic compositionsdescribed herein can also be used to sooth sensitive skin and to providesmooth and youthful skin with improved elasticity, reduced and delayedaging, enhanced youthful appearance and texture, and increasedflexibility, firmness, smoothness and suppleness.

[0059] The method of prevention and treatment according to thisinvention comprises administering topically to a host in need thereof atherapeutically effective amount of the formulated Free-B-Ringflavonoids and flavans isolated from a single source or multiplesources. The purity of the individual and/or a mixture of multipleFree-B-Ring flavonoids and flavans includes, but is not limited to 0.01%to 100%, depending on the methodology used to obtain the compound(s). Ina preferred embodiment, doses of the mixture of Free-B=Ring flavonoidsand flavans containing the same are an efficacious, nontoxic quantitygenerally selected from the range of 0.001% to 100% based on totalweight of the topical formulation. Persons skilled in the art usingroutine clinical testing are able to determine optimum doses for theparticular ailment being treated.

[0060] The present invention includes an evaluation of differentcompositions of Free-B-Ring flavonoids and flavans using enzymatic andin vivo models to optimize the formulation and obtain the desiredphysiological activity. The efficacy and safety of this formulation isalso demonstrated in human clinical studies. The compositions of thisinvention can be administered by any method known to one of ordinaryskill in the art. The modes of administration include, but are notlimited to, enteral (oral) administration, parenteral (intravenous,subcutaneous, and intramuscular) administration and topical application.In the preferred embodiment the method of treatment according to thisinvention comprises administering topically to a host in need thereof atherapeutically effective amount of a mixture of Free-B-Ring flavonoidsand flavans synthesized and/or isolated from a single plant or multipleplants.

[0061] It is to be understood that both the foregoing generaldescription and the following detailed description are exemplary andexplanatory only and are not restrictive of the invention as claimed.

BRIEF DESCRIPTION OF THE FIGURES

[0062]FIG. 1 depicts graphically a profile of the inhibition of COX-1and COX-2 by a standardized Free-B-Ring flavonoid extract (83% baicalinbased on HPLC) which was isolated from Scutellaria baicalensis. Theextract was examined for its inhibition of the peroxidase activity ofrecombinant ovine COX-1 (♦) and ovine COX-2 (▪). The data is presentedas percent inhibition vs. inhibitor concentration (μg/mL). The IC₅₀ forCOX-1 was calculated as 0.24 μg/mL/unit of enzyme while the IC₅₀ forCOX-2 was calculated as 0.48 μg/mL/unit.

[0063]FIG. 2 depicts graphically a profile of the inhibition of COX-1and COX-2 by the purified component baicalin which was isolated fromScutellaria baicalensis. The compound was examined for its inhibition ofthe peroxidase activity of recombinant ovine COX-1 (♦) and ovine COX-2(▪). The data is presented as percent inhibition vs. inhibitorconcentration (μg/mL). The IC₅₀ for COX-1 was determined to be 0.44μg/mL/unit of enzyme and the IC₅₀ for COX-2 was determined to be 0.28μg/mL/unit.

[0064]FIG. 3 depicts graphically a profile of the inhibition of COX-1and COX-2 by the purified component baicalein isolated from Scutellariabaicalensis. The compound was examined for its inhibition of theperoxidase activity of recombinant ovine COX-1 (♦) and ovine COX-2 (▪).The data is presented as percent inhibition vs. inhibitor concentration(μg/mL). The IC₅₀ for COX-1 was determined to be 0.18 μg/mL/unit ofenzyme and the IC₅₀ for COX-2 was determined to be 0.28 μg/mL/unit.

[0065]FIG. 4 depicts graphically a profile of the inhibition of COX-1and COX-2 by a standardized flavan extract containing 50% total flavanswhich was isolated from Acacia catechu. The extract was examined for itsinhibition of the peroxidase activity of recombinant ovine COX-1 (♦) andovine COX-2 (▪). The data is presented as percent inhibition vs.inhibitor concentration (μg/mL). The IC₅₀ for COX-1 was calculated as0.17 μg/mL/unit of enzyme and the IC₅₀ for COX-2 was calculated as 0.41μg/mL/unit.

[0066]FIG. 5 depicts graphically a profile of the inhibition of COX-1and COX-2 by the a composition of matter comprised of greater than 90%flavans isolated from Acacia catechu. The composition was examined forits inhibition of the peroxidase activity of recombinant ovine COX-1 (♦)and ovine COX-2 (▪). The data is presented as percent inhibition vs.inhibitor concentration (μg/mL). The IC₅₀ for COX-1 was calculated as0.11 μg/mL/unit of enzyme and the IC₅₀ for COX-2 was calculated as 0.42μg/mL/unit.

[0067]FIG. 6 depicts graphically a profile of the inhibition of COX-1and COX-2 by a formulation produced by combining an extract ofFree-B-Ring flavonoids isolated from the roots of Scutellariabaicalensis and an extract of flavans isolated from the bark of Acaciacatechu in a ratio of 80:20. This composition of matter, referred tohereinafter as Soliprin™, was examined for its inhibition of theperoxidase activity of recombinant ovine COX-1 (♦) and ovine COX-2 (▪).The data is presented as percent inhibition vs. inhibitor concentration(μg/mL). The IC₅₀ for COX-1 was calculated as 0.76 μg/mL/unit of enzymeand the IC₅₀ for COX-2 was calculated as 0.80 μg/mL/unit.

[0068]FIG. 7 depicts graphically a profile of the inhibition of COX-1and COX-2 by a formulation produced by combining an extract ofFree-B-Ring flavonoids isolated from the roots of Scutellariabaicalensis and an extract of flavans isolated from the bark of Acaciacatechu in a ratio of about 50:50. The composition, Soliprin™, wasexamined for its inhibition of the peroxidase activity of recombinantovine COX-1 (♦) and ovine COX-2 (▪). The data is presented as percentinhibition vs. inhibitor concentration (μg/mL). The IC₅₀ for COX-1 wascalculated as 0.38 μg/mL/unit of enzyme and the IC₅₀ for COX-2 wasdetermined to be 0.84 μg/mL/unit.

[0069]FIG. 8 depicts graphically a profile of the inhibition of COX-1and COX-2 by a formulation produced by combining an extract ofFree-B-Ring flavonoids isolated from the roots of Scutellariabaicalensis and an extract of flavans isolated from the bark of Acaciacatechu in a ratio of about 20:80. The composition, Soliprin™, wasexamined for its inhibition of the peroxidase activity of recombinantovine COX-1 (♦) and ovine COX-2 (▪). The data is presented as percentinhibition vs. inhibitor concentration (μg/mL). The IC₅₀ of thiscomposition for COX-1 was 0.18 μg/mL/unit of enzyme and the IC₅₀ forCOX-2 was 0.41 μg/mL/unit.

[0070]FIG. 9 depicts graphically a profile of the inhibition of 5-LO bythe flavan extract from Acacia catechu. The composition was examined forits inhibition of recombinant potato 5-lipoxygenase activity (♦) asdescribed in Example 4. The data is presented as percent inhibition ofassays without inhibitor. The IC₅₀ for 5-LO was 1.38 μg/mL/unit ofenzyme.

[0071]FIG. 10 illustrates the High Pressure Liquid Chromatography (HPLC)chromatogram of a typical formulation comprised of a mixture ofFree-B-Ring flavonoids isolated from the roots of Scutellariabaicalensis and flavans isolated from the bark of Acacia catechu in aratio of 80:20 carried out under the conditions as described in Example9.

[0072]FIG. 11 depicts graphically the effect of increasingconcentrations of Soliprin™ on the amount of LPS-induced newlysynthesized LTB₄ (♦) as determined by ELISA in THP-1 or HT-29 cells(ATCC) as described in Example 10. The Soliprin™ was produced throughthe combination of standardized extracts of Free-B-Ring flavonoidsisolated from the roots of Scutellaria baicalensis and flavans isolatedfrom the bark of Acacia catechu in a ratio of 80:20. The activity of theSoliprin™ formulation is expressed as % inhibition of induced LTB₄synthesis.

[0073]FIG. 12 compares the LTB₄ levels as determined by ELISA thatremain in HT-29 cells after treatment with 3 μg/mL Soliprin™ innon-induced cells to treatment with 3 μg/mL ibuprofen as described inExample 10. The Soliprin™ formulation demonstrated 80% inhibition ofLTB4 production in the HT-29 cells after two days of treatment.

[0074]FIG. 13 illustrates graphically ear-swelling data as a measure ofinhibition of inflammation as described in Example 11. Soliprin™produced through the combination of standardized extracts of Free-B-Ringflavonoids isolated from the roots of Scutellaria baicalensis andflavans isolated from the bark of Acacia catechu in a ratio of 80:20 wascompared to untreated mice and mice given indomethacin (1.5 μg/kg) viaoral gavage. The data is presented as the difference in micronmeasurement of the untreated vs. the treated ear lobe for each mouse.

[0075]FIG. 14 illustrates graphically the effect of 100 μg/kg ofSoliprin™, produced through the combination of standardized extracts ofFree-B-Ring flavonoids isolated from the roots of Scutellariabaicalensis and flavans isolated from the bark of Acacia catechu in aratio of 80:20 on the AA injected ankles of mice (Soliprin™+arachidonicacid) compared to non-treated mice (no treatment+arachidonic acid), micewithout AA injections (negative control) or mice that were injected withthe liquid carrier (vehicle control).

[0076]FIG. 15 depicts graphically the changes in hairless mice skinerythema scores in different treatment groups as a function of timefollowing irradiation of the mice with UV light as described in Example12. The mice in Groups B-1, A-1, B-2 and A-2 were treated with Soliprin™either before (Groups B-1 and B-2) or after (A-1 and A-2) irradiation.The Soliprin™ was produced through the combination of standardizedextracts of Free-B-Ring flavonoids isolated from the roots ofScutellaria baicalensis and flavans isolated from the bark of Acaciacatechu in a ratio of 80:20. With reference to FIG. 15, it can be seenthat topical applications of Soliprin™, both before and after UVradiation, significantly reduced erythema scores as compared with thecontrol group and the group that was administered the standard treatmentagent-Sooth-a-caine.

DETAILED DESCRIPTION OF THE INVENTION

[0077] Various terms are used herein to refer to aspects of the presentinvention. To aid in the clarification of the description of thecomponents of this invention, the following definitions are provided.

[0078] It is to be noted that the term “a” or “an” entity refers to oneor more of that entity; for example, a flavonoid refers to one or moreflavonoids. As such, the terms “a” or “an”, “one or more” and “at leastone” are used interchangeably herein.

[0079] “Free-B-Ring Flavonoids” as used herein are a specific class offlavonoids, which have no substitute groups on the aromatic B-ring, asillustrated by the following general structure:

[0080] wherein

[0081] R₁, R₂, R₃, R4, and R₅ are independently selected from the groupconsisting of —H, —OH, —SH, OR, —SR, —NH₂, —NHR, —NR₂, —NR₃ ⁺X⁻, acarbon, oxygen, nitrogen or sulfur, glycoside of a single or acombination of multiple sugars including, but not limited toaldopentoses, methyl-aldopentose, aldohexoses, ketohexose and theirchemical derivatives thereof;

[0082] wherein

[0083] R is an alkyl group having between 1-10 carbon atoms; and

[0084] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride, carbonate, etc.

[0085] “Flavans” as used herein refer to a specific class of flavonoids,which can be generally represented by the following general structure:

[0086] wherein

[0087] R₁, R₂, R₃, R₄ and R₅ are independently selected from the groupconsisting of H, —OH,

[0088] —SH, —OCH₃, —SCH₃, —OR, —SR, —NH₂, —NRH, —NR₂, —NR₃ ⁺X⁻, estersof substitution groups, including, but not limited to, gallate, acetate,cinnamoyl and hydroxyl-cinnamoyl esters, trihydroxybenzoyl esters andcaffeoyl esters and their chemical derivatives thereof; carbon, oxygen,nitrogen or sulfur glycoside of a single or a combination of multiplesugars including, but not limited to, aldopentoses, methyl aldopentose,aldohexoses, ketohexose and their chemical derivatives thereof; dimer,trimer and other polymerized flavans;

[0089] wherein

[0090] R is an alkyl group having between 1-10 carbon atoms; and

[0091] X is selected from the group of pharmaceutically acceptablecounter anions including, but not limited to hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride, carbonate, etc.

[0092] “Therapeutic” as used herein, includes treatment and/orprophylaxis. When used, therapeutic refers to humans as well as otheranimals.

[0093] “Pharmaceutically or therapeutically effective dose or amount”refers to a dosage level sufficient to induce a desired biologicalresult. That result may be the alleviation of the signs, symptoms orcauses of a disease or any other alteration of a biological system thatis desired.

[0094] “Placebo” refers to the substitution of the pharmaceutically ortherapeutically effective dose or amount dose sufficient to induce adesired biological that may alleviate the signs, symptoms or causes of adisease with a non-active substance.

[0095] A “host” or “patient” is a living subject, human or animal, intowhich the compositions described herein are administered. Thus, theinvention described herein may be used for veterinary as well as humanapplications and the terms “patient” or “host” should not be construedin a limiting manner. In the case of veterinary applications, the dosageranges can be determined as described below, taking into account thebody weight of the animal.

[0096] Note that throughout this application various citations areprovided. Each citation is specifically incorporated herein in itsentirety by reference.

[0097] The current invention provides methods for the extraction(Example 1, Table 1) of plants that contain Free-B-Ring flavonoids andflavans with organic and aqueous solvents. The crude extracts wereassayed for cyclooxygenase inhibitory activity (Example 2, Tables 2 and3). Purified Free-B-Ring flavonoids and flavans demonstrated inhibitoryactivity against cyclooxygenase (COX) and lipoxygenase (LOX),respectively, as shown in Examples 3 and 4. Methods for analyzing andquantifying the extracts are described in Examples 5 and 6 and theprocedures to generate standardized Free-B-Ring flavonoids and flavansfrom botanical origins are provided in Examples 7 and 8.

[0098] In one embodiment of the present invention, the standardizedFree-B-Ring flavonoid extract is comprised of the active compoundshaving a purity of between 1-99% (by weight) of total Free-B-Ringflavonoids as defined in Examples 1, 2, 5 and 8. Baicalin is the majoractive component in the extract, which accounts for approximately 50-90%(by weight) of the total Free-B-Ring flavonoids. In a preferredembodiment, the standardized extract contains >70% total Free-B-Ringflavonoids in which >75% of the Free-B-Ring flavonoids is baicalin.

[0099] In one embodiment, the standardized flavan extract is comprisedof the active compounds having a purity of between 1-99% (by weight)total flavans as defined in Examples 1, 4, 6 and 7. Catechin is themajor active component in the extract and accounts for 50-95% (byweight) of the total flavans. In a preferred embodiment, thestandardized flavan extract contains >80% total flavans in which >70% offlavans is catechin.

[0100] In one embodiment, Soliprin™ is produced by mixing the above twoextracts or synthetic compounds in a ratio from 99:1 to 1:99. Thepreferred ratios of Free-B-Ring flavonoids to flavans are 80:20 asdefined in Example 9 and Table 10 and 15:85 as defined in Example 9.

[0101] The concentration of Free-B-Ring flavonoids in Soliprin™ can befrom about 1% to 99% and the concentration of flavans in Soliprin™ canbe from 99% to 1%. In a preferred embodiment of the invention, theconcentration of total Free-B-Ring flavonoids in Soliprin™ isapproximately 20% with a baicalin content of approximately 15% of totalweight of the Soliprin™; and the concentration of total flavans inSoliprin™ is approximately 75% with a catechin content of approximately70%. In this embodiment, the total active components (Free-B-Ringflavonoids plus flavans) in Soliprin™ are >90% of the total weight.

[0102] The present invention includes methods that are effective insimultaneously inhibiting both the cyclooxygenase (COX) and lipoxygenase(LOX) enzymes, for use in the prevention and treatment of diseases andconditions related to the skin. The method for the simultaneous dualinhibition of the COX and LOX enzymes is comprised of administering,preferably topically a composition comprised of a mixture of Free-B-Ringflavonoids and flavans synthesized and/or isolated from a single plantor multiple plants to a host in need thereof. This composition of matteris referred to herein as Soliprin™. The efficacy of this method wasdemonstrated with purified enzymes, in different cell lines, in multipleanimal models and eventually in a human clinical study. The ratio of theFree-B-Ring flavonoids to flavans in the composition can be in the rangeof 99.9:0.1 of Free-B-Ring flavonoids:flavans to 0.1:99.9 Free-B-Ringflavonoids:flavans. In specific embodiments of the present invention,the ratio of Free-B-Ring flavonoids to flavans is selected from thegroup consisting of approximately 90:10, 80:20, 70:30, 60:40, 50:50,40:60, 30:70, 20:80 and 10:90. In a preferred embodiment of thisinvention, the ratio of Free-B-Ring flavonoids:flavans in thecomposition of matter is 20:80. In a preferred embodiment, theFree-B-Ring flavonoids are isolated from a plant or plants in theScutellaria genus of plants and the flavans are isolated from a plant orplants in the Acacia genus of plants.

[0103] The present invention also includes methods for the preventionand treatment of COX and LOX mediated diseases and conditions of theskin. The method for preventing and treating COX and LOX mediateddiseases and conditions of the skin is comprised of administering,preferably topically, to a host in need thereof an effective amount of acomposition comprised of a mixture of Free-B-Ring flavonoids and flavanssynthesized and/or isolated from a single plant or multiple plants and apharmaceutically acceptable carrier. The ratio of the Free-B-Ringflavonoids to flavans in the composition can be in the range of 99.9:0.1of Free-B-Ring flavonoids:flavans to 0.1:99.9 Free-B-Ringflavonoids:flavans. In specific embodiments of the present invention,the ratio of Free-B-ring flavonoids to flavans is selected from thegroup consisting of approximately 90:10, 80:20, 70:30, 60:40, 50:50,40:60, 30:70, 20:80 and 10:90. In a preferred embodiment of thisinvention, the ratio of Free-B-Ring flavonoids:flavans in thecomposition of matter is 20:80. In a preferred embodiment, theFree-B-Ring flavonoids are isolated from a plant or plants in theScutellaria genus of plants and the flavans are isolated from a plant orplants in the Acacia genus of plants.

[0104] In one embodiment, the present invention includes a method forpreventing and treating a number of COX and LOX mediated diseases andconditions of the skin including, but not limited to sun burns, thermalburns, acne, topical wounds, minor inflammatory conditions caused byfungal, microbial and viral infections, vitilago, systemic lupuserythromatosus, psoriasis, carcinoma, melanoma, as well as other mammalskin cancers. In another embodiment the present invention includes amethod for preventing and treating skin damage resulting from exposureto UV radiation, chemicals, heat, wind and dry environments. In yetanother embodiment the present invention includes a method forpreventing and treating wrinkles, saggy skin, lines and dark circlesaround the eyes, dermatitis and other allergy related conditions of theskin.

[0105] The present invention further includes therapeutic compositionscomprising the therapeutic agents of the present invention. In additionto their use for the prevention and treatment of the above describeddiseases and conditions of the skin, the therapeutic compositionsdescribed herein can be used to sooth sensitive skin and to providesmooth and youthful skin with improved elasticity, reduced and delayedaging, enhanced youthful appearance and texture, and increasedflexibility, firmness, smoothness and suppleness.

[0106] The Free-B-Ring flavonoids that can be used in accordance withthe instant invention include compounds illustrated by the generalstructure set forth above. The Free-B-Ring flavonoids of this inventionmay be obtained by synthetic methods or may be isolated from the familyof plants including, but not limited to Annonaceae, Asteraceae,Bignoniaceae, Combretaceae, Compositae, Euphorbiaceae, Labiatae,Lauranceae, Leguminosae, Moraceae, Pinaceae, Pteridaceae,Sinopteridaceae, Ulmaceae, and Zingiberacea. The Free-B-Ring flavonoidscan be extracted, concentrated, and purified from the following genus ofhigh plants, including but not limited to Desmos, Achyrocline, Oroxylum,Buchenavia, Anaphalis, Cotula, Gnaphalium, Helichrysum, Centaurea,Eupatorium, Baccharis, Sapium, Scutellaria, Molsa, Colebrookea, Stachys,Origanum, Ziziphora, Lindera, Actinodaphne, Acacia, Derris, Glycyrrhiza,Millettia, Pongamia, Tephrosia, Artocarpus, Ficus, Pityrogramma,Notholaena, Pinus, Ulmus and Alpinia.

[0107] The flavonoids can be found in different parts of plants,including but not limited to stems, stem barks, twigs, tubers, roots,root barks, young shoots, seeds, rhizomes, flowers and otherreproductive organs, leaves and other aerial parts. Methods for theisolation and purification of Free-B-Ring flavonoids are described inU.S. application Ser. No. 10/091,362, filed Mar. 1, 2002, entitled“Identification of Free-B-Ring Flavonoids as Potent Cox-2 Inhibitors,”which is incorporated herein by reference in its entirety.

[0108] The flavans that can be used in accordance with the method ofthis invention include compounds illustrated by the general structureset forth above. The flavans of this invention are isolated from a plantor plants selected from the Acacia genus of plants. In a preferredembodiment, the plant is selected from the group consisting of Acaciacatechu (A. catechu), A. concinna, A. farnesiana, A. Senegal, A.speciosa, A. arabica, A. caesia, A. pennata, A. sinuata. A. mearnsii, A.picnantha, A. dealbata, A. auriculiformis, A. holoserecia and A.mangium.

[0109] The flavans can be found in different parts of plants, includingbut not limited to stems, stem barks, trunks, trunk barks, twigs,tubers, roots, root barks, young shoots, seeds, rhizomes, flowers andother reproductive organs, leaves and other aerial parts. Methods forthe isolation and purification of flavans are described in U.S.application Ser. No. 10/104,477, filed Mar. 22, 2002, entitled“Isolation of a Dual Cox-2 and 5-Lipoxygenase Inhibitor from Acacia,”which is incorporated herein by reference in its entirety.

[0110] The present invention implements a strategy that combines aseries of in vivo inflammation and toxicity studies as well as in vitrobiochemical, cellular, and gene expression screens to identify activeplant extracts that specifically inhibit COX and LOX enzymatic activity,impact mRNA gene expression and reduce inflammation. The methods usedherein to identify active plant extracts that specifically inhibit COXand LOX are described in Examples 1 and 2, as well as in U.S.application Ser. No. 10/091,362, filed Mar. 1, 2002, entitled“Identification of Free-B-Ring Flavonoids as Potent Cox-2 Inhibitors;”U.S. application Ser. No. 10/104,477, filed Mar. 22, 2002, entitled“Isolation of a Dual Cox-2 and 5-Lipoxygenase Inhibitor from Acacia,”and U.S. application Ser. No. 10/427,746, filed Apr. 30, 2003, entitled“Formulation With Dual Cox-2 And 5-Lipoxygenase Inhibitory Activity,”each of which is incorporated herein by reference in its entirety.

[0111] The biochemical assay, used to measure inhibition of COX, relieson the protein's peroxidase activity in the presence of heme andarachidonic acid. This study which is described in Example 3, showedthat the purified Free-B-Ring flavonoids, baicalin and baicaleinisolated from Scutellaria baicalensis and the flavan extract isolatedfrom Acacia catechu, and each individual standardized extract containinghigh concentrations of Free-B-Ring flavonoids and flavans inhibited COXactivity (FIGS. 1-5). Additionally, compositions having different ratiosof each of the individual standardized extracts (i.e., 80:20, 50:50 and20:80 Free-B-Ring flavonoids:flavans), prepared as illustrated inExample 9, were all highly effective at inhibiting the COX activity invitro (FIGS. 6-8). The inhibition of LOX activity by a flavan extractisolated from Acacia catechu, was assessed using a lipoxygenasescreening assay in vitro as described in Example 4. The results areillustrated in FIG. 9. In addition, cell assays that targeted inhibitionof compounds in the breakdown of arachidonic acid in the LOX pathway,namely leukotriene B4 were performed using a Soliprin™ sample asdescribed in Example 10. The LTB₄ inhibition results by Soliprin™ areillustrated in FIGS. 11 and 12.

[0112] In vivo efficacy was demonstrated by the application of skinirritating substances, such as AA, to the ears and ankle joint of miceand measuring the reduction of swelling in mice treated with Soliprin™as described in Example 11. The results are set forth in FIGS. 13 and14. Finally, the efficacy of topical application of Soliprin™formulation in preventing and treating UV induced skin erythema isillustrated in Example 12 and FIG. 15. In the study described in Example12, Soliprin™ in a blend ratio of 80:20 as of Free-B-Ringflavonoids:flavans was dissolved in water and applied topically at twoconcentration to the skin of hairless mice both before and after UVexposure, respectively. The erythema scores of the hairless mice fromfour Soliprin™ groups, in both concentrations and regardless theapplications time as before or after UV exposure, all showed much lessredness in smaller skin areas as compared to severe and extendederythema in both the control group and the group that was treated withSooth-A Cain.

[0113] Example 13 (Tables 11 and 12) describes a general method for thepreparation of a Soliprin™ cream using pharmacologically,dermatologically and cosmetic acceptable excipients. For purposes ofillustration this Example provides a detailed procedure for thepreparation of both a 0.5 wt % and 1.5 wt % Soliprin™ cream. Finally,both of the Soliprin™ creams prepared as described in Example 13 wereevaluated on human skin for potential irritation and induction ofcontact sensitization. A total of 97 and 101 subjects completedinduction and challenge with the 0.5% and 1.5% Soliprin™ creams,respectively. Test results show that Soliprin™ creams at 0.5% and 1.5%concentration produced minimal irritation and did not elicit evidence ofinduced contact sensitization.

[0114] In summary, the present invention includes methods that areeffective in simultaneously inhibiting both the COX and LOX enzymes. Themethod for the simultaneous dual inhibition of the COX and LOX pathwaysis comprised of administering a composition comprising a mixture ofFree-B-Ring flavonoids and flavans synthesized and/or isolated from asingle plant or multiple plants to a host in need thereof. The ratio ofFree-B-Ring flavonoids to flavans in the composition can be in the rangeof 99:1 Free-B-Ring flavonoids:flavans to 1:99 of Free-B-Ringflavonoids:flavans. In specific embodiments of the present invention,the ratio of Free-B-Ring flavonoids to flavans is selected from thegroup consisting of approximately 90:10, 80:20, 70:30, 60:40, 50:50,40:60, 30:70,-20:80 and 10:90. In a preferred embodiment of theinvention, the ratio of Free-B-Ring flavonoids:flavans in thecomposition of matter is approximately 20:80. In a preferred embodiment,the Free-B-Ring flavonoids are isolated from a plant or plants in theScutellaria genus of plants and flavans are isolated from a plant orplants in the Acacia genus of plants.

[0115] The present further includes methods for the prevention andtreatment of COX and LOX mediated skin diseases and conditions. Themethod for preventing and treating COX and LOX mediated skin diseasesand conditions is comprised of administering to a host in need thereofan effective amount of a composition comprising a mixture of Free-B-Ringflavonoids and flavans synthesized and/or isolated from a single plantor multiple plants together with a pharmaceutically acceptable carrier.The ratio of Free-B-Ring flavonoids to flavans can be in the range of99:1 Free-B-Ring flavonoids:flavans to 1:99 of Free-B-Ringflavonoids:flavans. In specific embodiments of the present invention,the ratio of Free-B-Ring flavonoids to flavans is selected from thegroup consisting of approximately 90:10, 80:20, 70:30, 60:40, 50:50,40:60, 30:70, 20:80 and 10:90. In a preferred embodiment of theinvention, the ratio of Free-B-Ring flavonoids:flavans in thecomposition of matter is approximately 20:80. In a preferred embodiment,the Free-B-ring flavonoids are isolated from a plant or plants in theScutellaria genus of plants and flavans are isolated from a plant orplants in the Acacia genus of plants.

[0116] Applicant believes that U.S. application Ser. No. 10/104,477,filed Mar. 22, 2002, entitled “Isolation of a Dual COX-2 and5-Lipoxygenase Inhibitor from Acacia,” is the first report of acomposition of matter isolated from the Acacia genus of plants thatdemonstrates dual specificity for COX and LOX and that U.S. applicationSer. No. 10/091,362, filed Mar. 1, 2002, entitled “Identification ofFree-B-Ring Flavonoids as Potent COX-2 Inhibitors,” is the first reportof a correlation between Free-B-Ring flavonoid structure and COXinhibitory activity. These discoveries led to a novel blending of twoclasses of specific compounds—Free-B-Ring Flavonoids and flavans—to forma composition of matter, referred to herein as Soliprin™, which can beused for the prevention and treatment of COX and LOX mediated diseasesand conditions, as described in U.S. application Ser. No. 10/427,746,filed Apr. 30, 2003, entitled “Formulation With Dual Cox-2 And5-Lipoxygenase Inhibitory Activity.” COX and LOX mediated diseases andconditions include, but are not limited to diseases and conditions ofthe skin including, but are not limited to sun burns, thermal burns,acne, topical wounds, minor inflammatory conditions caused by fungal,microbial and viral infections, vitilago, systemic lupus erythromatosus,psoriasis, carcinoma, melanoma, as well as other mammal skin cancers,skin damage resulting from exposure to UV radiation, chemicals, heat,wind and dry environments, wrinkles, saggy skin, lines and dark circlesaround the eyes, dermatitis and other allergy related conditions of theskin. Although not limited by theory, it is believed that the mechanismof action of this class of compounds is the direct dual inhibition ofboth COX and LOX enzymatic activity.

[0117] The present invention further includes therapeutic compositionscomprising the therapeutic agents of the present invention includingvarious formulations thereof. Methods for the preparation of thesecompositions, together with methods for the determination of theirpurity and specific composition are described in Examples 5-9 and FIG.10.

[0118] In a preferred embodiment, the method of prevention and treatmentof COX and LOX mediated skin related diseases and conditions accordingto this invention comprises administering topically to a host in needthereof a therapeutically effective amount of the formulated Free-B-Ringflavonoids and/or flavans isolated from a single source or multiplesources. The purity of the individual and/or a mixture of Free-B-Ringflavonoids and flavans includes, but is not limited to 0.01% to 100%,depending on the methodology used to obtain the compound(s). In apreferred embodiment, doses of the mixture of Free-B-Ring flavonoidsand/or flavans containing that same are an efficacious, nontoxicquantity generally selected from the range of 0.001% to 100% based ontotal weight of the topical formulation. Persons skilled in the artusing routine clinical testing are able to determine optimum doses forthe particular ailment being treated.

[0119] The present invention includes evaluation of the differentcomposition of Free-B-Ring flavonoids and flavan using enzymatic and invivo anti-inflammation models to optimize the formulation and obtain thegreatest potency as described below. The present invention provides acommercially viable process for the isolation, purification andcombination of Acacia flavans with Free-B-Ring flavonoids to yield acomposition of matter having desirable physiological activity. Inaddition to their use for the prevention and treatment of the abovedescribed diseases and conditions of the skin, the therapeuticcompositions described herein can also be used to sooth sensitive skinand to provide smooth and youthful skin with improved elasticity,reduced and delayed aging, enhanced youthful appearance and texture, andincreased flexibility, firmness, smoothness and suppleness.

[0120] The compositions of the present invention can be formulated aspharmaceutical compositions which include other components such as apharmaceutically and/or cosmetically acceptable excipient, an adjuvant,and/or a carrier. For example, compositions of the present invention canbe formulated in an excipient that the host to be treated can tolerate.An excipient is an inert substance used as a diluent or vehicle for adrug. Examples of such excipients include, but are not limited to water,buffers, saline, Ringer's solution, dextrose solution, mannitol, Hank'ssolution, preservatives and other aqueous physiologically balanced saltsolutions. Nonaqueous vehicles, such as fixed oils, sesame oil, ethyloleate, or triglycerides may also be used. Other useful formulationsinclude suspensions containing viscosity enhancing agents, such assodium carboxymethylcellulose, sorbitol, or dextran. Excipients can alsocontain minor amounts of additives, such as substances that enhanceisotonicity and chemical stability. Examples of buffers includephosphate buffer, bicarbonate buffer, tris buffer, histidine, citrate,and glycine, or mixtures thereof, while examples of preservativesinclude, but are not limited to thimerosal, m- or o-cresol, formalin andbenzyl alcohol. Standard formulations can either be liquid or solids,which can be taken up in a suitable liquid as a suspension or solutionfor administration. Thus, in a non-liquid formulation, the excipient cancomprise dextrose, human serum albumin, preservatives, etc., to whichsterile water or saline can be added prior to administration.

[0121] In one embodiment of the present invention, the composition canalso include an adjuvant or a carrier. Adjuvants are typicallysubstances that generally enhance the biological response of a mammal toa specific bioactive agent. Suitable adjuvants include, but are notlimited to, Freund's adjuvant; other bacterial cell wall components;aluminum-based salts; calcium-based salts; silica; polynucleotides;toxoids; serum proteins; viral coat proteins; other bacterial-derivedpreparations; gamma interferon; block copolymer adjuvants, such asHunter's Titermax adjuvant (Vaxcel.TM., Inc. Norcross, Ga.); Ribiadjuvants (available from Ribi ImmunoChem Research, Inc., Hamilton,Mont.); and saponins and their derivatives, such as Quil A (availablefrom Superfos Biosector A/S, Denmark). Carriers are typically compoundsthat increase the half-life of a therapeutic composition in the treatedhost. Suitable carriers include, but are not limited to, polymericcontrolled release formulations, biodegradable implants, liposomes,bacteria, viruses, oils, esters, and glycols.

[0122] In one embodiment, the composition is prepared as a controlledrelease formulation, which slowly releases the composition of thepresent invention into the host. As used herein, a controlled releaseformulation comprises a composition of the present invention in acontrolled release vehicle. Suitable controlled release vehicles will beknown to those skilled in the art. Preferred controlled releaseformulations are biodegradable (i.e., bioerodible).

[0123] The therapeutic agents of the instant invention are preferablyadministered topically by any suitable means, known to those of skill inthe art for topically administering therapeutic compositions including,but not limited to as an ointment, gel, lotion, or cream base or as anemulsion, as a patch, dressing or mask, a nonsticking gauze, a bandage,a swab or a cloth wipe. Such topical application can be locallyadministered to any affected area, using any standard means known fortopical administration. A therapeutic composition can be administered ina variety of unit dosage forms depending upon the method ofadministration. For particular modes of delivery, a therapeuticcomposition of the present invention can be formulated in an excipientof the present invention. A therapeutic reagent of the present inventioncan be administered to any host, preferably to mammals, and morepreferably to humans. The particular mode of administration will dependon the condition to be treated.

[0124] In one embodiment, a suitable ointment is comprised of thedesired concentration of the mixture of Free-B-Ring flavonoids andflavans, that is an efficacious, nontoxic quantity generally selectedfrom the range of 0.001% to 100% based on total weight of the topicalformulation, from 65 to 100% (preferably 75 to 96%) of white softparaffin, from 0 to 15% of liquid paraffin, and from 0 to 7% (preferably3 to 7%) of lanolin or a derivative of synthetic equivalent thereof. Inanother embodiment the ointment may comprise a polyethylene—liquidparaffin matrix.

[0125] In one embodiment, a suitable cream is comprised of anemulsifying system together with the desired concentration of themixture of Free-B-Ring flavonoids and flavans as provided above. Theemulsifying system is preferably comprised of from 2 to 10% ofpolyoxyethylene alcohols (e.g. the mixture available under the trademarkCetomacrogol™ 1000), from 10 to 25% of stearyl alcohol, from 20 to 60%of liquid paraffin, and from 10 to 65% of water; together with one ormore preservatives, for example from 0.1 to 1% ofN,N″-methylenebis[N′-[3-(hydroxymethyl)-2,5-dioxo-4-imidazolidinyl]urea](available under the name Imidurea USNF), from 0.1 to 1% of alkyl4-hydroxybenzoates (for example the mixture available from NipaLaboratories under the trade mark Nipastat), from 0.01 to 0.1% of sodiumbutyl 4-hydroxybenzoate (available from Nipa Laboratories under thetrade mark Nipabutyl sodium), and from 0.1 to 2% of phenoxyethanol.Example 13 describes the formulation of two different concentrations ofthe composition of this invention as a cream and Example 14 describes astudy undertaken to evaluate the cream for irritation and sensitizationof the skin. From this study it was determined that Soliprin™ is a safecomposition that can be applied topically at an efficaciousconcentration without causing irritation or sensitization of the skin.

[0126] In one embodiment, a suitable gel is comprised of a semi-solidsystem in which a liquid phase is constrained within a three dimensionalpolymeric matrix with a high degree of cross-linking. The liquid phasemay be comprised of water, together with the desired amount of themixture of Free-B-Ring flavonoids and flavans, from 0 to 20% ofwater-miscible additives, for example glycerol, polyethylene glycol, orpropylene glycol, and from 0.1 to 10%, preferably from 0.5 to 2%, of athickening agent, which may be a natural product, for exampletragacanth, pectin, carrageen, agar and alginic acid, or a synthetic orsemi-synthetic compound, for example methylcellulose andcarboxypolymethylene (carbopol); together with one or morepreservatives, for example from 0.1 to 2% of methyl 4-hydroxybenzoate(methyl paraben) or phenoxyethanol-differential. Another suitable base,is comprised of the desired amount of the mixture of Free-B-Ringflavonoids and flavans, together with from 70 to 90% of polyethyleneglycol (for example, polyethylene glycol ointment containing 40% ofpolyethylene glycol 3350 and 60% of polyethylene glycol 400, prepared inaccordance with the U.S. National Formulary (USNF)), from 5 to 20% ofwater, from 0.02 to 0.25% of an anti-oxidant (for example butylatedhydroxytoluene), and from 0.005 to 0.1% of a chelating agent (forexample ethylenediamine tetraacetic acid (EDTA)).

[0127] The term soft paraffin as used above encompasses the cream orointment bases white soft paraffin and yellow soft paraffin. The termlanolin encompasses native wool fat and purified wool fat. Derivativesof lanolin include in particular lanolins which have been chemicallymodified in order to alter their physical or chemical properties andsynthetic equivalents of lanolin include in particular synthetic orsemisynthetic compounds and mixtures which are known and used in thepharmaceutical and cosmetic arts as alternatives to lanolin and may, forexample, be referred to as lanolin substitutes.

[0128] One suitable synthetic equivalent of lanolin that may be used isthe material available under the trademark Softisan™ known as Softisan649. Softisan 649, available from Dynamit Nobel Aktiengesellschaft, is aglycerine ester of natural vegetable fatty acids, of isostearic acid andof adipic acid; its properties are discussed by H. Hermsdorf in Fette,Seifen, Anstrichmittel, Issue No. 84, No.3 (1982), pp. 3-6.

[0129] The other substances mentioned hereinabove as constituents ofsuitable ointment or cream bases and their properties are discussed instandard reference works, for example pharmacopoeia. Cetomacrogol 1000has the formula CH₃(CH₂)_(m)(OCH₂CH₂)_(n)OH, wherein m may be 15 or 17and n may be 20 to 24. Butylated hydroxytoluene is2,6-di-tert-butyl-p-cresol. Nipastat is a mixture of methyl, ethyl,propyl and butyl 4-hydroxybenzoates.

[0130] The compositions of the invention may be produced by conventionalpharmaceutical techniques. Thus the aforementioned compositions, forexample, may conveniently be prepared by mixing together at an elevatedtemperature, preferably 60-70° C., the soft paraffin, liquid paraffin ifpresent, and lanolin or derivative or synthetic equivalent thereof. Themixture may then be cooled to room temperature, and, after addition ofthe hydrated crystalline calcium salt of mupirocin, together with thecorticosteroid and any other ingredients, stirred to ensure adequatedispersion.

[0131] Regardless of the manner of administration, the specific dose iscalculated according to the approximate body weight of the host. Furtherrefinement of the calculations necessary to determine the appropriatedosage for treatment involving each of the above mentioned formulationsis routinely made by those of ordinary skill in the art and is withinthe scope of tasks routinely performed by them without undueexperimentation, especially in light of the dosage information andassays disclosed herein. These dosages may be ascertained through use ofthe established assays for determining dosages utilized in conjunctionwith appropriate dose-response data.

[0132] It should be noted that the invention described herein may beused for veterinary as well as human applications and that the term“host” should not be construed in a limiting manner. In the case ofveterinary applications, the dosage ranges can be determined asdescribed above, taking into account the body weight of the animal.

[0133] The compositions of this invention can be administered by anymethod known to one of ordinary skill in the art. The modes ofadministration include, but are not limited to, enteral (oral)administration, parenteral (intravenous, subcutaneous, andintramuscular) administration and topical application. The method oftreatment according to this invention comprises administering internallyor topically to a patient in need thereof a therapeutically effectiveamount of a mixture of Free-B-Ring flavonoids and flavans synthesizedand/or isolated from a single plant or multiple plants. In a preferredembodiment the composition is administered topically.

[0134] The following examples are provided for illustrative purposesonly and are not intended to limit the scope of the invention.

EXAMPLES Example 1 Preparation of Organic and Aqueous Extracts fromAcacia and Scutellaria Plants

[0135] Plant material from Acacia catechu (L) Willd. barks, Scutellariaorthocalyx roots, Scutellaria baicalensis roots or Scutellarialateriflora whole plant was ground to a particle size of no larger than2 mm. Dried ground plant material (60 g) was then transferred to anErlenmeyer flask and methanol:dichloromethane (1:1) (600 mL) was added.The mixture was shaken for one hour, filtered and the biomass wasextracted again with methanol: dichloromethane (1:1) (600 mL). Theorganic extracts were combined and evaporated under vacuum to providethe organic extract (see Table 1 below). After organic extraction, thebiomass was air dried and extracted once with ultra pure water (600 mL).The aqueous solution was filtered and freeze-dried to provide theaqueous extract (see Table 1 below). TABLE 1 Yield of Organic andAqueous Extracts of Acacia and Scutellaria Species Plant Source AmountOrganic Extract Aqueous Extract Acacia catechu barks 60 g 27.2 g 10.8 gScutellaria orthocalyx roots 60 g 4.04 g 8.95 g Scutellaria baicalensis60 g 9.18 g 7.18 g roots Scutellaria lateriflora 60 g 6.54 g 4.08 g(whole plant)

Example 2 Inhibition of COX-2 and COX-1 Peroxidase Activity by PlantExtracts from Acacia catechu, Various Scutellaria Species and OtherPlants

[0136] The bioassay directed screening process for the identification ofspecific COX-2 inhibitors was designed to assay the peroxidase activityof the enzyme as described below.

[0137] Peroxidase Assay. The assay to detect inhibitors of COX-2 wasmodified for a high throughput platform (Raz). Briefly, recombinantovine COX-2 (Cayman) in peroxidase buffer (100 mM TBS, 5 mM EDTA, 1 μMHeme, 1 mg epinephrine, 0.094% phenol) was incubated with extract (1:500dilution) for 15 minutes. Quantablu (Pierce) substrate was added andallowed to develop for 45 minutes at 25° C. Luminescence was then readusing a Wallac Victor 2 plate reader. The results are presented in Table2.

[0138] Table 2 sets forth the inhibition of enzyme by the organic andaqueous extracts obtained from five plant species, including the bark ofAcacia catechu, roots of two Scutellaria species and extracts from threeother plant species, which are comprised of structurally similarFree-B-Ring flavonoids. Data is presented as the percent of peroxidaseactivity relative to the recombinant ovine COX-2 enzyme and substratealone. The percent inhibition by the organic extract ranged from 30% to90%. TABLE 2 Inhibition of COX-2 Peroxidase Activity by Various SpeciesInhibition of COX-2 Inhibition of COX-2 Plant Source by organic extractby aqueous extract Acacia catechu (bark) 75% 30% Scutellaria orthocalyx(root) 55% 77% Scutellaria baicalensis (root) 75%  0% Desmodiumsambuense 55% 39% (whole plant) Eucaluptus globulus (leaf) 30% 10%Murica nana (leaf) 90%  0%

[0139] Comparison of the relative inhibition of the COX-1 and COX-2isoforms requires the generation of IC₅₀ values for each of theseenzymes. The IC₅₀ is defined as the concentration at which 50%inhibition of enzyme activity in relation to the control is achieved bya particular inhibitor. In these experiments, IC₅₀ values were found torange from 6 to 50 μg/mL and 7 to 80 μg/mL for the COX-2 and COX-1enzymes, respectively, as set forth in Table 3. Comparison of the IC₅₀values of COX-2 and COX-1 demonstrates the specificity of the organicextracts from various plants for each of these enzymes. The organicextract of Scutellaria lateriflora for example, shows preferentialinhibition of COX-2 over COX-1 with IC₅₀ values of 30 and 80 μg/mL,respectively. While some extracts demonstrate preferential inhibition ofCOX-2, others do not. Examination of the HTP fractions and purifiedcompounds from these fractions is necessary to determine the truespecificity of inhibition for these extracts and compounds. TABLE 3 IC₅₀Values of Organic Extracts for Human and Ovine COX-2 and COX-1 IC₅₀Human IC₅₀ Ovine IC₅₀ Ovine COX-2 COX-2 COX-1 Plant Source (μg/mL)(μg/mL) (μg/mL) Acacia catechu (bark)  3 6.25 2.5 Scutellaria orthocalyx(root) Not done 10 10 Scutellaria baicalensis (root) 30 20 20Scutellaria lateriflora 20 30 80 (whole plant) Eucaluptus globulus(leaf) Not done 50 50 Murica nana (leaf)  5 6 7

Example 3 Inhibition of COX-1 and COX-2 Peroxidase Activity

[0140] In order to screen for compounds that inhibited the COX-1 andCOX-2 activities, a high throughput, in vitro assay was developed thatutilized the inhibition of the peroxidase activity of both enzymes.(Needleman et al. (1986) Annu Rev Biochem. 55:69). Briefly, thecomposition or compound being examined was titrated against a fixedamount of COX-1 and COX-2 enzymes. A cleavable, peroxide chromophore wasincluded in the assay to visualize the peroxidase activity of eachenzyme in presence of arachidonic acid as a cofactor. Typically, assayswere performed in a 96-well format. Each inhibitor, taken from a 10mg/mL stock solution in 100% DMSO, was tested in triplicate at roomtemperature using the following range of concentrations: 0, 0.1, 1, 5,10, 20, 50, 100, and 500 μg/mL. To each well, 150 μL of 100 mM Tris-HCl,pH 7.5 was added along with 10 μL of 22 μM Hematin diluted in trisbuffer, 10 μL of inhibitor diluted in DMSO and 25 units of either theCOX-1 or COX-2 enzyme. The components were mixed for 10 seconds on arotating platform, followed by the addition of 20 μL of 2 mMN,N,N′N′-tetramethyl-p-phenylenediamine dihydrochloride (TMPD) and 20 μLof 1.1 mM arachidonic acid to initiate the reaction. The plate wasshaken for 10 seconds and then incubated 5 minutes before reading theabsorbance at 570 nm. The inhibitor concentration vs. % inhibition wasplotted and the IC₅₀ determined by taking the half-maximal point alongthe isotherm and intersecting the concentration on the X-axis. The IC₅₀was then normalized to the number of enzyme units in the assay. Theresults are summarized in Table 4. TABLE 4 Inhibition of COX EnzymeActivity by Purified Free-B-Ring Flavonoids Free-B-Ring FlavonoidsInhibition of COX-1 Inhibition of COX-2 Baicalein 107%  109% 5,6-Dihydroxy-7- 75% 59% methoxyflavone 7,8-Dihydroxyflavone 74% 63%Baicalin 95% 97% Wogonin 16% 12%

[0141] The dose responses and IC₅₀ values for a standardized Free-B-Ringflavonoid extract, baicalin, and baicalein isolated from the roots ofScutellaria baicalensis are provided in FIGS. 1, 2 and 3, respectively.The dose responses and IC₅₀ values for two standardized flavan extract(50% and >90% flavans, respectively) isolated from the heartwood ofAcacia catechu are provided in FIGS. 4 and 5, respectively. The doseresponses and IC₅₀ values for three formulations of Free-B-Ringflavonoids and flavans of varying composition are provided in FIG. 6(80:20 blending), FIG. 7 (50:50 blending) and FIG. 8 (20:80 blending),respectively.

Example 4 Inhibition of 5-Lipoxygenase by Catechin isolated from Acaciacatechu

[0142] One of the most important pathways involved in the inflammatoryresponse is produced by non-heme, iron-containing lipoxygenases (5-LO,12-LO, and 15-LO), which catalyze the addition of molecular oxygen ontofatty acids such as AA (AA) to produce the hydroperoxides 5-, 12- and15-HPETE, which are then converted to leukotrienes. There were earlyindications that the flavan extract from A. catechu may provide somedegree of LOX inhibition, thereby preventing the formation of 5-HPETE. ALipoxygenase Inhibitor Screening Assay Kit (Cayman Chemical, Inc., Cat#760700) was used to assess whether an extract isolated from A. catechucontaining >90% flavans directly inhibited LOX in vitro. The 15-LO fromsoybeans normally used in the kit was replaced with potato LOX, after abuffer change from phosphate to a tris -based buffer usingmicrofiltration was performed. This assay detects the formation ofhydroperoxides through an oxygen sensing chromagen. Briefly, the assaywas performed in triplicate by adding 90 μL of 0.17 units/μL potato5-LO, 20 μL of 1.1 mM AA, 100 μL of oxygen-sensing chromagen and 10 μLof purified flavan inhibitor to final concentrations ranging from 0 to500 μg/mL. The IC₅₀ for 5-LO inhibition from this composition wasdetermined to be 1.38 μg/mL/unit of enzyme. The results are set forth inFIG. 9.

Example 5 HPLC Quantification of Free-B-Ring Flavonoids in ActiveExtracts Isolated from Scutellaria orthocalyx (Roots) Scutellariabaicalensis (Roots) and Oroxylum indicum (Seeds)

[0143] The presence and quantity of Free-B-Ring flavonoids in fiveactive extracts isolated from three different plant species as describedin Examples 1 and 2 were determined by HPLC and the results are setforth in the Table 5, below. The Free-B-Ring flavonoids werequantitatively analyzed by HPLC on a Luna C-18 column (250×4.5 mm, 5 μm)using a 1% phosphoric acid and acetonitrile gradient from 80% to 20% in22 minutes. The Free-B-Ring flavonoids were detected using a UV detectorat 254 nm and identified based on retention time by comparison withbaicalin, baicalein and other Free-B-Ring flavonoid standards. TABLE 5Free-B-Ring Flavonoid Content in Active Plant Extracts % Total %Extractible amount of Free-B-Ring Weight from Free-B-Ring FlavonoidsActive Extracts of Extract BioMass Flavonoids in Extract S. orthocalyx8.95 g 14.9%  0.2 mg 0.6% (aqueous extract) S. orthocalyx 3.43 g 5.7%1.95 mg 6.4% (organic extract) S. baicalensis 7.18 g 12.0% 0.03 mg0.07%  (aqueous extract) S. baicalensis 9.18 g 15.3% 20.3 mg 35.5% (organic extract) Oroxylum indicum 6.58 g 11.0%  0.4 mg 2.2% (organicextract)

Example 6 HPLC Quantification of Active Extracts from Acacia catechu

[0144] The flavans in the organic and aqueous extracts isolated fromAcacia catechu as illustrated in Examples 1 and 2 were quantified byHPLC using a PhotoDiode Array detector (HPLC/PDA) and a Luna C18 column(250 mm×4.6 mm). The flavans were eluted from the column using anacetonitrile gradient from 10% to 30% ACN over a period of 20 minutes,followed by 60% ACN for five minutes. The results are set forth in Table6. The flavans were quantified based on retention time and PDA datausing catechin and epicatechin as standards. The retention times for thetwo major flavans were 12.73 minutes and 15.76 minutes, respectively.TABLE 6 Free-B-Ring Flavonoid Content in Active Plant Extracts ActiveExtracts from Weight of % Extractible % Flavans bark of A. catechuExtract from BioMass in Extract Aqueous Extract 10.8 g 18.0% 0.998%Organic Extract 27.2 g 45.3% 30.37%

Example 7 Preparation of a Standardized Extract from Acacia catechu

[0145]Acacia catechu (500 mg of ground root) was extracted twice with 25mL (2×25 mL) of the following solvent systems. (1) 100% water, (2) 80:20water:methanol, (3) 60:40 water:methanol, (4) 40:60 water:methanol, (5)20:80 water:methanol, (6) 100% methanol, (7) 80:20 methanol:THF, (8)60:40 methanol:THF. The two extracts from each individual extractionwere combined concentrated and dried under low vacuum. Theidentification of the chemical components in each extract was achievedby HPLC using a PhotoDiode Array detector (HPLC/PDA) and a 250 mm×4.6 mmC18 column. The chemical components were quantified based on retentiontime and PDA data using catechin and epicatechin as standards. Theresults are set forth in Table 7. As shown in Table 7, the flavanextract generated from solvent extraction with 80% methanol/waterprovided the highest concentration of flavan components. TABLE 7Solvents for Generating Standardized Flavan Extracts from Acacia catechuTotal Extraction Weight of % Extractible amount of % Catechins SolventExtract from BioMass Catechins in Extract 100% water 292.8 mg 58.56% 13mg 12.02% water:methanol 282.9 mg 56.58% 13 mg 11.19% (80:20)water:methanol 287.6 mg 57.52% 15 mg 13.54% (60:40) water:methanol 264.8mg 52.96% 19 mg 13.70% (40:60) water:methanol 222.8 mg 44.56% 15 mg14.83% (20:80) 100% methanol 215.0 mg 43.00% 15 mg 12.73% methanol:THF264.4 mg 52.88% 11 mg 8.81% (80:20) methanol:THF 259.9 mg 51.98% 15 mg9.05% (60:40)

[0146] Higher purity material can be obtained by recrystallization ofextracts having a catechin content of between 8%-15% using analcohol/water and/or aqueous solvents as the recrystallization solvent.It may be necessary to decolorize prior to recrystallization by addingactive charcoal or other decolorization agent to a heated saturatedsolution of the extract. The high purity catechins then crystallizedupon cooling of the heated saturated solution. The crystals were thenfiltered to remove solvent, dried and ground into a fine powder.Recrystallization can be repeated as necessary to achieve a the desiredlevel of purity (60%-100% of catechin flavans).

Example 8 Preparation of Standardized Free-B-Ring Flavonoid Extractsfrom Various Scutellaria species

[0147]Scutellaria orthocalyx (500 mg of ground root) was extracted twicewith 25 mL of the following solvent systems. (1) 100% water, (2) 80:20water:methanol, (3) 60:40 water:methanol, (4) 40:60 water:methanol, (5)20:80 water:methanol, (6) 100% methanol, (7) 80:20 methanol:THF, (8)60:40 methanol:THF. The extracts were combined, concentrated and driedunder low vacuum. Identification of chemical components in each extractwas performed by HPLC using a PhotoDiode Array detector (HPLC/PDA) and a250 mm×4.6 mm C18 column. The chemical components were quantified basedon retention time and PDA data using baicalein, baicalin, scutellarein,and wogonin as standards. The results are set forth in Table 8. TABLE 8Quantification of Free-B-Ring Flavonoids Extracted from Scutellariaorthocalyx % Extractible Total % Extraction Weight of from amount ofFlavonoids in Solvent Extract BioMass Flavonoids Extract 100% water   96mg 19.2% 0.02 mg 0.20% Water:methanol 138.3 mg 27.7% 0.38 mg 0.38%(80:20) Water:methanol 169.5 mg 33.9% 0.78 mg 8.39% (60:40)Water:methanol 142.2 mg 28.4% 1.14 mg 11.26% (40:60) Water:methanol104.5 mg 20.9% 0.94 mg 7.99% (20:80) 100% methanol  57.5 mg 11.5% 0.99mg 10.42% methanol:THF  59.6 mg 11.9% 0.89 mg 8.76% (80:20) methanol:THF 58.8 mg 11.8% 1.10 mg 10.71% (60:40)

[0148]Scutellaria baicalensis (1000 mg of ground root) was extractedtwice using 50 mL of a mixture of methanol and water as follows: (1)100% water, (2) 70:30 water:methanol, (3) 50:50 water:methanol, (4)30:70 water:methanol, (5) 100% methanol. The extracts were combined,concentrated and dried under low vacuum. Identification of the chemicalcomponents was performed by HPLC using a PhotoDiode Array detector(HPLC/PDA), and a 250 mm×4.6 mm C18 column. The chemical components ineach extract were quantified based on retention time and PDA data usingbaicalein, baicalin, scutellarein, and wogonin standards. The resultsare set forth in Table 9. TABLE 9 Quantification of Free-B-RingFlavonoids Extracted from Scutellaria baicalensis % Extractible Total %Extraction Weight of from amount of Flavonoids in Solvent ExtractBioMass Flavonoids Extract 100% water 277.5 mg 27.8%   1 mg  0.09%Water:methanol 338.6 mg 33.9% 1.19 mg 11.48% (70:30) Water:methanol304.3 mg 30.4% 1.99 mg 18.93% (50:50) Water:methanol 293.9 mg 29.4% 2.29mg 19.61% (30:70) 100% methanol 204.2 mg 20.4% 2.73 mg 24.51%

[0149] Higher purity Free-B-Ring flavonoids can be obtained byrecrystallization of extracts having a Free-B-Ring flavonoid content ofbetween 8-15% using alcohol/water as a recrystallization solvent. It maybe necessary to decolorize prior to recrystallization by adding activecharcoal or other decolorization agent to a heated saturated solution ofthe extract. The Free-B-Ring flavonoids crystallized upon cooling. Thecrystals were filtered, dried and ground into a fine powder.Recrystallization can be repeated as necessary to achieve a the desiredlevel of purity (60%-100% of Free-B-Ring flavonoids).

Example 9 Preparation of a Formulation with a Standardized Free-B-RingFlavonoid Extract from the Roots of Scutellaria baicalensis and aStandardized Flavan Extract from the Bark of Acacia catechu

[0150] A novel composition of matter, referred to herein as Soliprin™was formulated using two standardized extracts isolated from Acacia andScutellaria, respectively, together with one or more excipients. Ageneral example for preparing such a composition is set forth below. TheAcacia extract used in this example contained >80% total flavans, ascatechin and epicatechin, and the Scutellaria extract contained >80%Free-B-Ring flavonoids, which was primarily baicalin. The Scutellariaextract also contained other minor amounts of Free-B-Ring flavonoids asset forth in Table 11. One or more excipients/preservatives was alsoadded to the composition of matter. The ratio of flavans and Free-B-Ringflavonoids can be adjusted based on the indications and the specificrequirements with respect to inhibition of COX vs. LO, requirements ofskin penetration, and potency requirements of the product, such asduration of potency required, etc. The quantity of the excipients can beadjusted based on the actual active content of each ingredient. Ablending table for each individual batch of product must be generatedbased on the product specification and QC results for individual batchof ingredients. Additional amounts of active ingredients in the range of2-5% are recommended to meet the product specification.

[0151]Scutellaria baicalensis root extract (38.5 kg) (lot #RM052302-01)having a Free-B-Ring flavonoid content of 82.2% (baicalin); Acaciacatechu bark extract (6.9 kg) (lot #RM052902-01) with total flavancontent of 80.4%; and excipient (5.0 kg of Candex) were combined toprovide a Soliprin™ formulation (50.4 kg) having a blending ratio of85:15 by weight of the active Free-B-Ring flavonoids and flavans. Table10 provides the quantification of the active Free-B-Ring flavonoids andflavans of this specific batch of Soliprin™ (Lot #G1702-COX-2),determined using the methods provided in Examples 6 and 8. Withreference to Table 10, this specific batch of Soliprin™ contains 86%total active ingredients, including 75.7% Free-B-Ring flavonoids and10.3% flavans. FIG. 10 illustrates the HPLC chromatogram of arepresentative Soliprin™ sample which had a blending ratio of 80:20 byweight of the active Free-B-Ring flavonoids and flavans. TABLE 10Free-B-Ring Flavonoid and Flavan Content of a Soliprin ™ FormulationActive Components % Content 1. Flavonoids a. Baicalin 62.5% b. MinorFlavonoids  i. Wogonin-7-glucuronide  6.7%  ii. Oroxylin A 7-glucuronide 2.0%  iii. Baicalein  1.5%  iv. Wogonin  1.1%  v. Chrysin-7-glucuronide 0.8%  vi. 5-Methyl-wogonin-7-glucuronide  0.5%  vii. Scutellarin  0.3% viii. Norwogonin  0.3%  ix. Chrysin <0.2%  x. Oroxylin A <0.2% c. TotalFree-B-ring Flavonoids 75.7% 2. Flavans a. Catechin  9.9% b. Epicatechin 0.4% c. Subtotal Flavans 10.3% 3. Total Active Ingredients   86%

[0152] Using the same approach, the following batches of Soliprin™ wereprepared using a combination of a standardized Free-B-Ring flavonoidextract from Scutellaria baicalensis roots and a standardized flavanextract from Acacia catechu bark having a blending ratio of 12:88 and15:85, respectively.

[0153]Scutellaria baicalensis root extract (58.0 g) (lot #RM021203-01)having a Free-B-Ring flavonoid content of 87.9% (as baicalin) and Acaciacatechu bark extract (442.0 g) (lot #RM050603-01) with total flavancontent of 84.9% were blended to provide a Soliprin™ composition (500 g,lot #QJ205-19) having a blending ratio of 12:88 by weight. Utilizing themethods provided in Examples 6 and 8, the Free-B-Ring flavonoid contentof (baicalin) was 9.65% and flavan content (total catechin andepicatechin) was 73.2% in this specific batch of Soliprin™ (lot#QJ205-19).

[0154]Scutellaria baicalensis root extract (300 g) (lot #RM060403-01)having a Free-B-Ring flavonoid content of 82.9% (as baicalin) and Acaciacatechu bark extract (1700 g) (lot #RM050603-01) with total flavancontent of 90.8% were blended to provide a Soliprin™ composition (2000g, lot #A1904) having a blending ratio of 15:85 by weight. Utilizing themethods provided in Examples 6 and 8, the Free-B-Ring flavonoid content(baicalin) was 15.6% and flavan content (total catechin and epicatechin)was 75.0% in this specific batch of Soliprin™ (lot #A1904).

Example 10 Measurements of Dose Response and IC₅₀ Values of 5-LO EnzymeInhibition from a Formulation of Soliprin™

[0155] A Soliprin™ formulation (80:20) was prepared as described inExample 9. (See also Example 14 of U.S. patent application Ser. No.10/427,746, filed Apr. 30, 2003, entitled “Formulation With Dual COX-2And 5-Lipoxygenase Inhibitory Activity,” which is incorporated herein byreference in its entirety) using a combination of a standardizedFree-B-Ring flavonoid extract from Scutellaria baicalensis roots and astandardized flavan extract from Acacia catechu bark with a blendingratio of 80:20. The sample was titrated in tissue culture mediacontaining THP-1 or HT-29 cells; monocyte cell lines that express COX-1,COX-2 and 5-LOX. A competitive ELISA for Leukotriene B4 (LTB4; Neogen,Inc., Cat#406110) was used to assess the effect of this Soliprin™formulation on newly synthesized levels of LTB4 present in each cellline as a measure Soliprin™'s inhibitory effect on the 5-LOX pathway.The assay was performed in duplicate by adding 160,000 to 180,000 cellsper well in 6-well plates. The Soliprin™ formulation was added to theTHP-1 cultures at 3, 10, 30 and 100 μg/mL and incubated overnight(˜12-15 hrs) at 37° C. with 5% CO₂ in a humidified environment. Theresults are set forth in FIG. 11, which shows that the production ofnewly LPS-induced LTB4 was almost completely inhibited by the additionof Soliprin™ to the THP-1 cultures between 3 and 10 μg/mL.

[0156] Soliprin™ and ibuprofen, another known 5-LOX inhibitor, wereadded to the HT-29 cells at 3 μg/mL and incubated 48 hrs at 37° C. with5% CO₂ in a humidified environment. Each treated cell line was thenharvested by centrifugation and disrupted by gentle douncehomogenization lysis in physiological buffers. As shown in FIG. 12,Soliprin™ inhibited generation of 80% of the newly synthesized LTB4 inHT-29 cells. Ibuprofen only showed a 20% reduction in the amount of LTB4over the same time period.

Example 11 Evaluation of the Efficacy of Soliprin™ with in vivo MouseEar Swelling Model

[0157] A Soliprin™ formulation was prepared using a combination of astandardized Free-B-Ring flavonoid extract from Scutellaria baicalensisroots and a standardized flavan extract from Acacia catechu bark with ablending ratio of 80:20 as described in Example 9. To test whether thiscomposition could be used to treat inflammation in vivo, the compositionwas administered by oral gavage to 4-5 week old ICR mice (Harlan Labs)one day before treatment of their ears with arachidonic acid (AA). Testmice were fed dose equivalents of 50, 100 and 200 mg/kg of Soliprin™suspended in olive oil while control mice were fed only olive oil. Thefollowing day, 20 μL of 330 mM AA in 95% alcohol was applied to one earof each mouse, while alcohol was applied to the other ear as a control.Mice treated with Soliprin™ showed a measurable dose response thattracked with increasing doses of Soliprin™, as demonstrated in FIG. 13.With reference to FIG. 13, the 200 mg/kg dose reduces swelling by over50% as compared to the “No treatment” control. The 50 mg/kg dose ofSoliprin™ was as effective as the 50 mg/kg dose of another stronganti-inflammatory, indomethacin.

[0158] In another animal model designed to demonstrate theanti-inflammatory activity of Soliprin™ the 80:20 formulation describedabove was orally administrated to mice in a dose of 100 mg/kg suspendedin olive oil ˜12 hours before injection of 20 μL of 100 mM AA in 95%ethanol into the hind ankle joints of 4-5 week old ICR mice (HarlanLabs). The test group was fed the Soliprin™ formulation, while anothergroup was not given the formulation. Control groups included mice thathad not received arachidonic acid injections (negative control) and agroup that had 95% ethanol without AA injected (vehicle control). Thesegroups were also not given Soliprin™. The results are set forth in FIG.14. With reference to FIG. 14, the mice given Soliprin™ that wereinjected with AA showed background levels of swelling as compared to thecontrols and the untreated arachidonic injected group. These resultsdemonstrate the effectiveness of Soliprin™ for reducing swelling injoints, the site of action.

Example 12 Evaluation of the Efficacy of Soliprin™ in Preventing andTreating Damage Resulting from Exposure of Skin to UV Radiation

[0159] Six groups of hairless female mice (five mice per group) (StrainSKH-1, Harlan Labs) were irradiated, while anesthetized, for threeminutes on three consecutive days with 0.626 mW/cm² to test theeffectiveness of the Soliprin™ formulation in preventing and treatingdamage resulting from exposure of skin to UV radiation. The Soliprin™formulation was prepared using a combination of a standardizedFree-B-Ring flavonoid extract from Scutellaria baicalensis roots and astandardized flavan extract from Acacia catechu bark with a blendingratio of 80:20 as described in Example 9. The six treatment groups wereas follows: Group # 1 Control group: no treatment before or after UVirradiation 2 Positive control: treated with a topical application ofSooth-A-Caine (Banana Boat) after UV irradiation 3 Soliprin ™ TreatmentB-1: treated with topical application of 1 mg/mL Soliprin ™ in waterbefore UV irradiation 4 Soliprin ™ Treatment A-1: treated with topicalapplication of 1 mg/mL Soliprin ™ in water after UV irradiation 5Soliprin ™ Treatment B-2: treated with topical application of 5 mg/mLSoliprin ™ in water before UV irradiation 6 Soliprin ™ Treatment A-2:treated with topical application of 5 mg/mL Soliprin ™ in water after UVirradiation

[0160] After three days of UV exposure and treatment, the mice werescored on level of erythema (redness) using the following scale: 0—novisible erythema; 1—very slight erythema; 2—well defined erythema;3—severe erythema; and 4—tumor formation. Erythema was scored by eye foreach group. The results are set forth in FIG. 15. With reference to FIG.15 it can be seen that the control group (Group 1) had severe redness onday 3 (72 hours after the three day exposure to UV radiation). TheSooth-a-caine group also had maximum redness on day 3 (Group 2). Theredness for the Soliprin™ treated groups (Groups 3-6) never exceeded ascore of 2. These scores, though subjective, show that Soliprin™ iseffective in both preventing and treating UV caused skin erythema.

[0161] Photographs of representative mice on day four clearlydemonstrate differences between the control group, the Sooth-a-cain™treated groups and the Soliprin™ treated groups (data not shown). Thecontrol group and Sooth-a-cain™ treated animals exhibited very extensivepatterns and redness of erythema compared to the animals treated withthe Soliprin™ formulation both before and after UV exposure. The animalstreated before UV irradiation with 5 mg/mL Soliprin™ exhibited the leastamount of erythema as compared to all of the other animals.

Example 13 Formulation of the Soliprin™ Composition into a Cream

[0162] Two different concentrations of Soliprin™ (0.5% and 1.5% byweight of Soliprin™) (lot #A1904 as described in Example 9) wereformulated as creams as illustrated in the following procedures and inTables 11 and 12.

[0163] Soliprin™ (Lot #A1904) was dissolved in water at room temperatureand homogenized with a blender until it was fully dispersed in solution(approximately 5 minutes). At room temperature and without stirring oragitating the solution, Ultrez-21 carbomer was added by sprinkling ontothe surface of the solution and allowing it to fully wet (no white areasvisible) and fall into the solution. With gentle stirring, the solutionwas then heated to 40° C. and glycerin was added (Part A). The mixturewas then stirred for an additional 5 minutes. The remaining components(Part B) were weighed and heated to 40° C. while mixing. At 40° C., theremaining components (Part B) were added to Part A and the resultingcomposition was mixed well until homogenous (approximately 5 minutes).The emulsion was cooled to 30° C. and the pH was adjusted toapproximately 5.5 (5.3 to 5.7) by titrating with neutralizer whilestirring with a stir bar and/or spatula. The emulsion became highlyviscous due to neutralization-induced conformational change of thecarbomer. The emulsion eventually achieved a suitable viscosity for anemulsion cream. The emulsion cream was then mixed until uniform afterwhich it was poured into a clean storage vessel and stored at 2° to 8°C. for one month. TABLE 11 Ingredient list for a 0.5% Soliprin CreamPhase Ingredient % (w/w) Weight (g) Aqueous Water, Purified 85.00 1275.0Soliprin (Lot#A1904) 0.50 7.5 Ultrez 21 Carbomer 0.50 7.5 Glycerin 8.00120.0 Oil PEG-7 Glyceryl Cocoate 3.00 45.0 Caprylic/Capric Triglyceride2.67 40.0 PH Sodium Hydroxide (18% w/v), 0.00 0.0 Neutralizer MolecularBiology Grade SUM 7 Ingredients 99.7 1495.0

[0164] TABLE 12 Ingredient list in a 1.5% Soliprin Cream PhaseIngredient % (w/w) Weight (g) Aqueous Water, Purified 84.00 1260.0Soliprin (Lot#A1904) 1.50 22.5 Ultrez 21 Carbomer 0.50 7.5 Glycerin 8.00120.0 Oil PEG-7 Glyceryl Cocoate 3.00 45.0 Caprylic/Capric Triglyceride2.67 40.0 pH Sodium Hydroxide (18% w/v), Neutralizer Molecular BiologyGrade SUM 7 Ingredients 99.7 1495.0

Example 14 Evaluation of a Soliprin™ Cream for Irritation and Inductionof Contact Sensitization by Repetitive Application to Human Skin

[0165] The Soliprin™ was tested on human skin using an adaptation of theDraize Patch Test (Marzulli and Maibach (1977) Contact Allergy:Predictive Testing in Humans. In Advances in Modem Toxicology,Dermatotoxicology and Pharmacology. Eds. Marzulli, F. N and Maibach, H.I. 4, 353-372). The test sites were located on the upper arm or theparaspinal region of the back. Each test article had an induction siteand a challenge site. The induction site was comprised of two sub-sites:an original-site and a move-site. Patches, which contains 0.2 ml ofSoliprin cream on each patch, were applied repeatedly to theoriginal-site unless a sufficiently strong irritation reactiondeveloped, requiring the patch to be applied to the move-site. Patcheswere applied by a clinical research institute and were removed anddiscarded by the subjects approximately 24 or 48/72 hours later. In theinduction phase, repetitive application of the test article to the samesite on the skin and a total of 9 induction patches were applied withina 4-week period. The rest period was 10 to 21 days between applicationof the last induction patch and application of the challenge patch.During this time no test article or any other material was applied tothe test area. At the challenge phase, the test article was applied to anaive site on the opposite side of the body and discarded by thesubjects approximately 24 or 48 hours later.

[0166] Skin responses to each patch application were examined and gradedunder light supplied by a 100-watt incandescent blue bulb according tothe designated scoring scale. In instances where a strong irritationreaction warranted application of the test article to the move-site,residual scores were be recorded through the end of induction (or untilresolved if reactions persist after induction is completed) for allpreviously exposed sites. All skin reactions were recorded. During thechallenge phase, skin responses were evaluated approximately 48 and 72or 96 hours after patch application. Conclusions, with regard to inducedsensitivity, were derived primarily from the challenge evaluations.

[0167] The two Soliprin™ creams prepared in the Example 13 at 0.5% and1.5% Soliprin™ concentrations were evaluated according to the aboveprotocol. A total of 120 subjects were recruited for each group.Ninety-seven subjects completed the study for the 0.5% Soliprin™ groupand 101 subjects completed the study for 1.5% Soliprin™ group. There wasno evidence of sensitization reaction for either the 0.5% and 1.5%Soliprin™ creams. For the 0.5% Soliprin™, during induction, sixteensubjects exhibited occasional occurrences of slight to mild erythema(scores of + and/or 1). At challenge, four subjects exhibited slight tomild erythema at 48 hours that cleared by 96 hours. For 1.5% Soliprin™,during induction, twenty-six subjects exhibited occasional occurrencesof slight to mild erythema (scores of + and/or 1). At challenge, onesubjects exhibited slight to mild erythema at 48 hours that cleared by96 hours.

[0168] This study demonstrates that Soliprin™ is a safe ingredient thatcan be applied topically to human skin at an efficacious concentrationwithout causing irritation or sensitization.

1. A method for preventing and treating cyclooxygenase (COX) andlipoxygenase (LOX) mediated diseases and conditions of the skin, saidmethod comprising administering to a host in need thereof an effectiveamount of a pharmaceutical composition comprising a mixture of at leastone Free-B-ring flavonoid and at least one flavan.
 2. The method ofclaim 1 wherein the ratio of Free-B-Ring flavonoid to flavan in saidcomposition is selected from the range of 99:1 Free-B-Ringflavonoid:flavan to 1:99 of Free-B-Ring flavonoid:flavan.
 3. The methodof claim 2 wherein the ratio of Free-B-Ring flavonoid:flavan in thecomposition of matter is about 20:80.
 4. The method of claim 1 whereinsaid Free-B-Ring flavonoid is selected from the group of compoundshaving the following structure:

wherein R₁, R₂, R₃, R₄, and R₅ are independently selected from the groupconsisting of —H, —OH, —SH, —OR, —SR, —NH₂, —NHR, —NR₂, —NR₃ ⁺X⁻, acarbon, oxygen, nitrogen or sulfur, glycoside of a single or acombination of multiple sugars including, aldopentoses,methyl-aldopentose, aldohexoses, ketohexose and their chemicalderivatives thereof, wherein R is an alkyl group having between 1-10carbon atoms; and X is selected from the group of pharmaceuticallyacceptable counter anions including, hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride and carbonate:
 5. The method ofclaim 1 wherein said flavan is selected from the group of compoundshaving the following structure:

wherein R₁, R₂, R₃, R₄ and R₅ are independently selected from the groupconsisting of H, —OH, —SH, —OCH₃, —SCH₃, —OR, —SR, —NH₂, —NRH, —NR₂,—NR₃ ⁺X⁻, esters of substitution groups, independently selected from thegroup consisting of gallate, acetate, cinnamoyl and hydroxyl-cinnamoylesters, trihydroxybenzoyl esters and caffeoyl esters; a carbon, oxygen,nitrogen or sulfur glycoside of a single or a combination of multiplesugars including, aldopentoses, methyl aldopentose, aldohexoses,ketohexose and their chemical derivatives thereof; dimer, trimer andother polymerized flavans; wherein R is an alkyl group having between1-10 carbon atoms; and X is selected from the group of pharmaceuticallyacceptable counter anions including, but not limited to hydroxyl,chloride, iodide, sulfate, phosphate, acetate, fluoride, carbonate. 6.The method of claim 1 wherein said Free-B-Ring flavonoid and said flavanare obtained by organic synthesis or are isolated from a plant.
 7. Themethod of claim 6 wherein said Free-B-Ring flavonoid and said flavan areisolated from a plant part selected from the group consisting of stems,stem barks, trunks, trunk barks, twigs, tubers, roots, root barks, youngshoots, seeds, rhizomes, flowers and other reproductive organs, leavesand other aerial parts.
 8. The method of claim 6 wherein saidFree-B-Ring flavonoid is isolated from a plant family selected from thegroup consisting of Annonaceae, Asteraceae, Bignoniaceae, Combretaceae,Compositae, Euphorbiaceae, Labiatae, Lauranceae, Leguminosae, Moraceae,Pinaceae, Pteridaceae, Sinopteridaceae, Ulmaceae and Zingiberacea. 9.The method of claim 6 wherein said Free-B-Ring flavonoid is isolatedfrom a plant genus selected from the group consisting of Desmos,Achyrocline, Oroxylum, Buchenavia, Anaphalis, Cotula, Gnaphalium,Helichrysum, Centaurea, Eupatorium, Baccharis, Sapium, Scutellaria,Molsa, Colebrookea, Stachys, Origanum, Ziziphora, Lindera, Actinodaphne,Acacia, Derris, Glycyrrhiza, Millettia, Pongamia, Tephrosia, Artocarpus,Ficus, Pityrogramma, Notholaena, Pinus, Ulmus and Alpinia.
 10. Themethod claim 6 wherein said flavan is are isolated from a plant speciesselected from the group consisting of the Acacia catechu, Acaciaconcinna, Acacia farnesiana, Acacia Senegal, Acacia speciosa, Acaciaarabica, A. caesia, A. pennata, A. sinuata. A. mearnsii, A. picnantha,A. dealbata, A. auriculiformis, A. holoserecia and A. mangium.
 11. Themethod of claim 6 wherein said Free-B-ring flavonoid is isolated from aplant or plants in the Scutellaria genus of plants and said flavan isisolated from a plant or plants in the Acacia genus of plants.
 12. Themethod of claim 1 wherein the composition is administered in a dosageselected from 0.001 to 200 mg/kg of body weight.
 13. The method of claim1 wherein the composition is administered in a pharmaceutical,dermatological or cosmetic formulation comprised of approximately 0.001weight percent (wt %) to 40.0 wt % of the mixture of Free-B-Ringflavonoids and flavans in a pharmaceutically, dermatologically andcosmetically acceptable carrier.
 14. The method of claim 1 wherein theroutes of the administration are selected from the group consisting oftopical, aerosol, suppository, intradermic, intramusclar, andintravenous administration.
 15. The method of claim 14 wherein the routeof the administration is topical.
 16. The method of claim 15 wherein thecomposition is administered using a nonsticking gauze, a bandage, aswab, a cloth wipe, a patch, a mask, a protectant, a cleanser, anantiseptic, a solution, a cream, a lotion, an ointment, a gel or anemulsion, a liquid, a paste, a soap, or a powder.
 17. The method ofclaim 1 wherein the pharmaceutical composition is further comprised of aconventional excipient that is pharmaceutically, dermatologically andcosmetically suitable for topical application and optionally anadjuvant, and/or a carrier, and/or a regular or controlled releasingvehicle.
 18. The method of claim wherein the COX and LOX mediateddiseases and conditions of the skin are selected from the groupconsisting of sun burns, thermal burns, acne, topical wounds, minorinflammatory conditions caused by fungal, microbial and viralinfections, vitilago, systemic lupus erythromatosus, psoriasis,carcinoma, melanoma, as well as other mammal skin cancers, skin damageresulting from exposure to ultraviolet (UV) radiation, chemicals, heat,wind and dry environments, wrinkles, saggy skin, lines and dark circlesaround the eyes, dermatitis and other allergy related conditions of theskin.
 19. A pharmaceutical composition of matter for use in theprevention and treatment of diseases and conditions related to the skincomprised of a mixture of at least one Free-B-ring flavonoid and atleast one flavan.
 20. The pharmaceutical composition of claim 19 whereinthe ratio of Free-B-Ring flavonoid to flavan in said composition isselected from the range of 99:1 Free-B-Ring flavonoid:flavan to 1:99 ofFree-B-Ring flavonoid:flavan.
 21. The pharmaceutical composition of 20wherein the ratio of Free-B-Ring flavonoid:flavan in the composition ofmatter is about 20:80.
 22. The pharmaceutical composition of claim 19wherein said Free-B-Ring flavonoid is selected from the group ofcompounds having the following structure:

wherein R₁, R₂, R₃, R₄, and R₅ are independently selected from the groupconsisting of —H, —OH, —SH, —OR, —SR, —NH₂, —NHR, —NR₂, —NR₃ ⁺X⁻, acarbon, oxygen, nitrogen or sulfur, glycoside of a single or acombination of multiple sugars including, aldopentoses,methyl-aldopentose, aldohexoses, ketohexose and their chemicalderivatives thereof; wherein R is an alkyl group having between 1-10carbon atoms; and X is selected from the group of pharmaceuticallyacceptable counter anions including, hydroxyl, chloride, iodide,sulfate, phosphate, acetate, fluoride and carbonate.
 23. Thepharmaceutical composition of claim 19 wherein said flavan is selectedfrom the group of compounds having the following structure:

wherein R₁, R₂, R₃, R₄ and R₅ are independently selected from the groupconsisting of H, —OH, —SH, —OCH₃, —SCH₃, —OR, —SR, —NH₂, —NRH, —NR₂,—NR₃ ⁺X⁻, esters of substitution groups, independently selected from thegroup consisting of gallate, acetate, cinnamoyl and hydroxyl-cinnamoylesters, trihydroxybenzoyl esters and caffeoyl esters; a carbon, oxygen,nitrogen or sulfur glycoside of a single or a combination of multiplesugars including, aldopentoses, methyl aldopentose, aldohexoses,ketohexose and their chemical derivatives thereof; dimer, trimer andother polymerized flavans; wherein R is an alkyl group having between1-10 carbon atoms; and X is selected from the group of pharmaceuticallyacceptable counter anions including, but not limited to hydroxyl,chloride, iodide, sulfate, phosphate, acetate, fluoride, carbonate. 24.The pharmaceutical composition of claim 19 wherein said Free-B-Ringflavonoid and said flavan are obtained by organic synthesis or areisolated from a plant.
 25. The pharmaceutical composition of claim 25wherein said Free-B-Ring flavonoid and said flavan are isolated from aplant part selected from the group consisting of stems, stem barks,trunks, trunk barks, twigs, tubers, roots, root barks, young shoots,seeds, rhizomes, flowers and other reproductive organs, leaves and otheraerial parts.
 26. The pharmaceutical composition of claim 25 whereinsaid Free-B-Ring flavonoid is isolated from a plant family selected fromthe group consisting of Annonaceae, Asteraceae, Bignoniaceae,Combretaceae, Compositae, Euphorbiaceae, Labiatae, Lauranceae,Leguminosae, Moraceae, Pinaceae, Pteridaceae, Sinopteridaceae, Ulmaceaeand Zingiberacea.
 27. The pharmaceutical composition of claim 25 whereinsaid Free-B-Ring flavonoid is isolated from a plant genus selected fromthe group consisting of Desmos, Achyrocline, Oroxylum, Buchenavia,Anaphalis, Cotula, Gnaphalium, Helichrysum, Centaurea, Eupatorium,Baccharis, Sapium, Scutellaria, Molsa, Colebrookea, Stachys, Origanum,Ziziphora, Lindera, Actinodaphne, Acacia, Derris, Glycyrrhiza,Millettia, Pongamia, Tephrosia, Artocarpus, Ficus, Pityrogramma,Notholaena, Pinus, Ulmus and Alpinia.
 28. The pharmaceutical compositionof claim 25 wherein said flavan is are isolated from a plant speciesselected from the group consisting of the Acacia catechu, Acaciaconcinna, Acacia farnesiana, Acacia Senegal, Acacia speciosa, Acaciaarabica, A. caesia, A. pennata, A. sinuata. A. mearnsii, A. picnantha,A. dealbata, A. auriculiformis, A. holoserecia and A. mangium.
 29. Thepharmaceutical composition of claim 25 wherein said Free-B-ringflavonoid is isolated from a plant or plants in the Scutellaria genus ofplants and said flavan is isolated from a plant or plants in the Acaciagenus of plants.
 30. The pharmaceutical composition of claim 19 furthercomprising a pharmaceutically acceptable excipient and optionally anadjuvant or a carrier.
 31. The pharmaceutical composition of claim 19,wherein said composition is formulated for topical application.
 32. Thepharmaceutical composition of claim 19 where said composition isformulated in a regular or controlled releasing vehicle.
 33. A methodfor simultaneously inhibiting the enzymatic activity of the COX and LOXenzymes in the skin comprised of administering to a host in need thereofan effective amount of a pharmaceutical composition comprised of amixture of at least one Free-B-Ring flavonoids and one flavan.
 34. Themethod of claim 34 wherein the pharmaceutical composition isadministered in a pharmaceutical, dermatological or cosmetic formulationcomprised of approximately 0.001 weight percent (wt %) to 40.0 wt % ofthe mixture of Free-B-Ring flavonoids and flavans in a pharmaceutically,dermatologically and cosmetically acceptable carrier.
 35. The method ofclaim 34 wherein the routes of the administration are selected from thegroup consisting of topical, aerosol, suppository, intradermic,intramusclar, and intravenous administration.
 36. The method of claim 36wherein the route of the administration is topical.
 37. The method ofclaim 34 wherein the pharmaceutical composition is administered using anonsticking gauze, a bandage, a swab, a cloth wipe, a patch, a mask, aprotectant, a cleanser, an antiseptic, a solution, a cream, a lotion, anointment, a gel or an emulsion, a liquid, a paste, a soap, or a powder.38. The method of claim 34 wherein the pharmaceutical composition isfurther comprised of a conventional excipient that is pharmaceutically,dermatologically and cosmetically suitable for topical application andoptionally an adjuvant, and/or a carrier, and/or a regular or controlledreleasing vehicle.
 39. A method for improving mammal skin appearancemediated by COX and LOX pathways comprising administering to a host inneed thereof an effective amount of a composition comprising a mixtureof at least one Free-B-Ring flavonoid and at least one flavan.
 40. Themethod of claim 40 wherein the pharmaceutical composition isadministered in a pharmaceutical, dermatological or cosmetic formulationcomprised of approximately 0.001 weight percent (wt %) to 40.0 wt % ofthe mixture of Free-B-Ring flavonoids and flavans in a pharmaceutically,dermatologically and cosmetically acceptable carrier.
 41. The method ofclaim 40 wherein the routes of the administration are selected from thegroup consisting of topical, aerosol, suppository, intradermic,intramusclar, and intravenous administration.
 42. The method of claim 42wherein the route of the administration is topical.
 43. The method ofclaim 40 wherein the pharmaceutical composition is administered using anonsticking gauze, a bandage, a swab, a cloth wipe, a patch, a mask, aprotectant, a cleanser, an antiseptic, a solution, a cream, a lotion, anointment, a gel or an emulsion, a liquid, a paste, a soap, or a powder.44. The method of claim 40 wherein the pharmaceutical composition isfurther comprised of a conventional excipient that is pharmaceutically,dermatologically and cosmetically suitable for topical application andoptionally an adjuvant, and/or a carrier, and/or a regular or controlledreleasing vehicle.